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AU - Burge BW, Pfefferkorn ER
TI - Isolation and characterization of conditional-lethal mutants of Sindbis virus.
SO - Virology 1966 Oct;30(2):204-13

AU - Burge BW, Pfefferkorn ER
TI - Complementation between temperature-sensitive mutants of Sindbis virus.
SO - Virology 1966 Oct;30(2):214-23

AU - Burge BW, Pfefferkorn ER
TI - Temperature-sensitive mutants of Sindbis virus: biochemical correlates of complementation.
SO - J Virol 1967 Oct;1(5):956-62

AU - Yin FH, Lockart RZ Jr
TI - Maturation defects in temperature-sensitive mutants of Sindbis virus.
SO - J Virol 1968 Jul;2(7):728-37

AU - Burge BW, Pfefferkorn ER
TI - Functional defects of temperature-sensitive mutants of Sindbis virus.
SO - J Mol Biol 1968 Jul 14;35(1):193-205

AU - Ben-Ishai Z, Goldblum N, Becker Y
TI - The intracellular site and sequence of Sindbis virus replication.
SO - J Gen Virol 1968 May;2(3):365-75

AU - Lab M, Tinland R, Kirn A
TI - [Influence of temperature on the development of Sindbis virus and on the production of interferon in cell cultures]. [French]
SO - Can J Microbiol 1968 Nov;14(11):1211-6

AU - Scheele CM, Pfefferkorn ER
TI - Inhibition of interjacent ribonucleic acid (26S) synthesis in cells infected by Sindbis virus.
SO - J Virol 1969 Aug;4(2):117-22

AU - Dobos P, Faulkner P
TI - Characterization of a cytoplasmic fraction from Sindbis virus-infected cultures.
SO - Can J Microbiol 1969 Feb;15(2):215-22

AU - Sreevalsan T, Yin FH
TI - Sindbis virus-induced viral ribonucleic acid polymerase.
SO - J Virol 1969 Jun;3(6):599-604

AU - Strauss JH Jr, Burge BW, Darnell JE
TI - Sindbis virus infection of chick and hamster cells: synthesis of virus-specific proteins.
SO - Virology 1969 Mar;37(3):367-76

AU - Mussgay M, Enzmann PJ, Horst J
TI - Influence of an arbovirus infection (Sindbis virus) on the protein and ribonucleic acid synthesis of cultivated chick embryo cells.
SO - Arch Gesamte Virusforsch 1970;31(1):81-92

AU - Scheele CM, Pfefferkorn ER
TI - Virus-specific proteins synthesized in cells infected with RNA+ temperature-sensitive mutants of Sindbis virus.
SO - J Virol 1970 Mar;5(3):329-37

AU - Kitsak VIa
TI - [Changes in the hereditary properties of Sindbis virus in long-term cultivation under conditions of lowered temperature]. [RUSSIAN]
SO - Vopr Virusol 1970 Mar-Apr;15(2):177-81

AU - Nicoli J, Coulomb P, Meyer F
TI - [Structural proteins of Sindbis virus and translation of the viral genome]. [French]
SO - Ann Inst Pasteur (Paris) 1971 May;120(5):683-97

AU - Cheban DS, Kitsak VIa
TI - [A change in various biological properties of Sindbis virus as a result of prolonged passage at low temperatures]. [RUSSIAN]
SO - Vopr Virusol 1972 Jul-Aug;17(4):451-3

AU - Stollar V, Shenk TE
TI - Homologous viral interference in Aedes albopictus cultures chronically infected with Sindbis virus.
SO - J Virol 1973 Apr;11(4):592-5

AU - Rasmussen LE, Armstrong JA, Ho M
TI - Interference mediated by a variant of Sindbis virus. II. Actinomycin D-sensitive interference in the absence of interferon.
SO - J Infect Dis 1973 Aug;128(2):163-9

AU - Rasmussen LE, Armstrong JA, Ho M
TI - Interference mediated by a variant of Sindbis virus. I. Isolation of an avirulent variant and in vivo interference.
SO - J Infect Dis 1973 Aug;128(2):156-62

AU - Waite MR
TI - Protein synthesis directed by an RNA temperature-sensitive mutant of Sindbis virus.
SO - J Virol 1973 Feb;11(2):198-206

AU - Snyder HW, Sreevalsan T
TI - Proteins specified by Sindbis virus in chick embryo fibroblast cells.
SO - Biochem Biophys Res Commun 1973 Jul 2;53(1):24-31

AU - Bras-Herreng F
TI - [Changes in the properties of Sindbis virus during passage experiments in "Drosophila" (author's transl)]. [French]
SO - Ann Microbiol (Paris) 1973 Jun;124A(4):507-33

AU - Poiree J, Bonnet J, Negre G, Nicoli J
TI - [Polyribosomes and ribonucleic acids associated with polyribosomes in cells infected by a togavirus, the Sindbis virus]. [French]
SO - Ann Microbiol (Paris) 1973 Oct;124(3):371-85

AU - Zhdanov VM, Azadova NB, Kopel'man RN
TI - [The biophysical characteristics of viral structures in chronic Sindbis infection]. [RUSSIAN]
SO - Vopr Virusol 1973 Sep-Oct;18(5):524-8

AU - Schlesinger S, Weiss B, Goran D, Schlesinger M, Cancedda R
TI - Formation of RNA and protein in cells infected with standard and defective Sindbis virus.
SO - Med Microbiol Immunol (Berl) 1974;160(4):311-29

AU - Peleg J, Stollar V
TI - Homologous interference in Aedes aegypti cell cultures infected with Sindbis virus.
SO - Arch Gesamte Virusforsch 1974;45(4):309-18

AU - Atkins GJ, Samuels J, Kennedy SI
TI - Isolation and preliminary characterization of temperature-sensitive mutants of Sindbis virus strain AR339.
SO - J Gen Virol 1974 Dec;25(3):371-80

AU - Shenk TE, Koshelnyk KA, Stollar V
TI - Temperature-sensitive virus from Aedes albopictus cells chronically infected with Sindbis virus.
SO - J Virol 1974 Feb;13(2):439-47

AU - Snyder HW, Sreevalsan T
TI - Proteins specified by Sindbis virus in HeLa cells.
SO - J Virol 1974 Feb;13(2):541-4

AU - Sreevalsan T, Rosemond-Hornbeak H
TI - Inhibition of Sindbis virus replication in HeLa cells by poliovirus.
SO - Antimicrob Agents Chemother 1974 Jan;5(1):55-62

AU - Waite MR, Lubin M, Jones KJ, Bose HR
TI - Phosphorylated proteins of Sindbis virus.
SO - J Virol 1974 Jan;13(1):244-6

AU - Hunt JM, Marcus PI
TI - Mechanism of Sindbis virus-induced intrinsic interference with vesicular stomatitis virus replication.
SO - J Virol 1974 Jul;14(1):99-109

AU - Segal S, Sreevalsan T
TI - Sindbis virus replicative intermediates: purification and characterization.
SO - Virology 1974 Jun;59(2):428-42

AU - Raul F, Egly JM, Kempf J
TI - Characterization of a particulate replicative structure in sindbis virus infected cells.
SO - FEBS Lett 1974 Jun 15;42(3):314-8

AU - Simmons DT, Strauss JH
TI - Translation of Sindbis virus 26 S RNA and 49 S RNA in lysates of rabbit reticulocytes.
SO - J Mol Biol 1974 Jun 25;86(2):397-409

AU - Johnston RE, Wan K, Bose HR
TI - Homologous interference induced by Sindbis virus.
SO - J Virol 1974 Nov;14(5):1076-82

AU - Simmons DT, Strauss JH
TI - Replication of Sindbis virus. V. Polyribosomes and mRNA in infected cells.
SO - J Virol 1974 Sep;14(3):552-9

AU - Cancedda R, Swanson R, Schlesinger MJ
TI - Viral proteins formed in a cell-free rabbit reticulocyte system programmed with RNA from a temperature-sensitive mutant of Sindbis virus.
SO - J Virol 1974 Sep;14(3):664-71

AU - Chudzio T, Inglot AD
TI - Biological properties of three spontaneous mutants of Sindbis virus.
SO - Arch Immunol Ther Exp (Warsz) 1975;23(1):113-29

AU - Bras-Herreng F
TI - [Multiplication of sindbis virus in Drosophila cells cultivated in vitro (author's transl)]. [French]
SO - Arch Virol 1975;48(2):121-9
AB - Sindbis virus replicates in Drosophila cell cultures without any cytopathic effect. In continuous cell lines the virus is able to establish a persistent infection similar to other arboviruses in insect cell lines. The growth curve of Sindbis virus in Drosophila cells shows a maximum virus yield at around 24 hours postinfection, thereafter the virus production decreases and then remains fairly constant during a number of cell divisions. The average virus yield per cell has been estimated to be small, i.e. at best 5 PFU per day.

AU - Stollar V, Shenk TE, Koo R, Igarashi A, Schlesinger RW
TI - Observations of Aedes albopictus cell cultures persistently infected with Sindbis virus.
SO - Ann N Y Acad Sci 1975;266:214-31

AU - Peleg J
TI - In vivo behavior of a Sindbis virus mutant isolated from persistently infected Aedes aegypti cell cultures.
SO - Ann N Y Acad Sci 1975;266:204-13
AB - A mutant of the Sindbis virus SV-S was found to interfere with the regular course of infection by the wild strain of the virus SV-W in A. aegypti mosquitoes and in suckling mice. In mosquitoes, this result was manifested by a reduced titer of SV-W in the presence of SV-S and by a failure of the mosquitoes to transmit SV-W. In the brains of suckling mice, in the presence of SV-S, the growth of sc inoculated SV-W was suppressed, and as a result, the usually lethal course of infection by this virus was converted into a nonlethal one.

AU - Kos KA, Osborne BA, Goldsby RA
TI - Inhibition of group B arbovirus antigen production and replication in cells enucleated with cytochalasin B.
SO - J Virol 1975 Apr;15(4):913-7
AB - A comparative study of the growth of Sindbis (SIN) virus, a group A arbovirus (togavirus), and Japanese encephalitis (JE) virus, a representative group B arbovirus (togavirus), was conducted in enucleate and nucleate cells. Immunofluorescent tests and yield measurements demonstrated that chicken embryo cells which had been enucleated and subsequently infected with SIN virus produced virus-specific antigens and infectious virus. By contrast, JE failed to replicate or produce virus-specific antigen in cells which had been enucleated before or even 2 h post infection. Studies of the effect of enulceation at various times after infection demonstrated that a nucleus must be present at least 2 and possibly as long as 4 h after infection to produce either JE-specific antigen or infectious JE virus. These studies demonstrate that the replication of SIN, a group A arbovirus (togavirus), which has no nuclear requirement, contrasts sharply with that of a group B arbovirus (togavirus), JE, which may have an initial dependence on a nucleus-associated process.

AU - Kaluza G, Kraus AA, Rott R
TI - Inhibition of cellular protein synthesis by simultaneous pretreatment of host cells with fowl plague virus and actinomycin D: a method for studying early protein synthesis of several RNA viruses.
SO - J Virol 1975 Jan;17(1):1-9
AB - A method is described for analysis of viral protein synthesis early after infection when minute amounts of viral proteins are effectively concealed by large amounts of produced host-specific proteins. The method is superior to a radioimmune assay, since all virus-induced proteins can be measured independent of their immunological reactivity. Host-specific protein synthesis can be suppressed by infection with fowl plague virus. Addition of actinomycin C 1.25 h postinfection does not prevent this suppression, but it does block effectively the formation of fowl plague virus-specific proteins. Such cells synthesize only small amounts of cellular proteins, as revealed by polyacrylamide electrophoresis. They can be superinfected with several different enveloped viruses, however, without significant diminution of virus yeilds. In pretreated cells the eclipse is shortened for Semliki Forest virus, Sindbis virus, and vesicular stomatitis virus, but prolonged for Newcastle disease virus. The onset of protein synthesis, specific for the superinfecting virus, could be clearly demonstrated within 1 h after superinfection. At this time, in cells superinfected with Semliki Forest virus, great amounts of NSP 75 (nonstructural protein; molecular weight, 75 X 10(3)) and reduced amounts of the core protein C could be deomonstrated. The precursor glycoprotein NSP 68 is followed by a new polypeptide, NSP 65: three proteins with molecular weights exceeding 100 X 10(3) were observed which are missing later in the infectious cycle. Similar results were obtained after superinfection with Sindbis virus. The formation of a new polypeptide with a molecular weight of about 80 X 10(3) was detected. After superinfection with vesicular stomatis virus or Newcastle disease virus the formation of new proteins, characteristic for the early stage of infeciton, was not observed.

AU - Schlesinger MJ
TI - Function of Sindbis virus 49S and 26S RNAs in in vitro protein synthesizing systems. A summary.
SO - Med Biol 1975 Oct;53(5):380-2

AU - Cancedda R, Villa-Komaroff L, Lodish HF, Schlesinger M
TI - Initiation sites for translation of sindbis virus 42S and 26S messenger RNAs.
SO - Cell 1975 Oct;6(2):215-22
AB - Sindbis virus 26S RNA is the principal species of virus-specific RNA found in the infected cell; it is derived from a one third segment of virion 42S RNA. When translated in cell-free extracts from mouse ascites cells or rabbit reticulocytes, 26S RNA directed the synthesis primarily of the 33,000 dalton virus capsid protein, and the protein products were in the form of free peptides rather than peptidyl-tRNA. In contrast, the polypeptides synthesized in either extract in response to Sindbis virus 42S RNA were heterogeneous, ranging in molecular weight from 33,000 to 190,000, and were largely in the form of peptidyl-tRNA. The number of independent initiation sites on the 26S and 42S RNAs was determined by analyzing a tryptic digest of reaction products labeled with yeast N-formyl-35S-methionyl-tRNAFmet. The 26S RNA appeared to contain a single initiation site, and this site could also be found in varying amounts in different preparations of 42S RNA. However, a second initiation site, distinct from that of 26S RNA, was the major site in 42S virion RNA. These results suggest that 42S virion RNA contains two potential sites for initiation of protein synthesis. Only one of these may be active, however, and it is postulated that the second site functions primarily, if not exclusively, in the subgenomic 26S RNA species. In this regard, Sindbis virus 42S RNA may represent a novel form of a eucaryotic messenger RNA.

AU - Schlesinger RW
TI - Sindbis virus replication in vertebrate and mosquito cells: an interpretation. [Review]
SO - Med Biol 1975 Oct;53(5):295-301
AB - This paper summarizes recent comparative studies of Sindbis virus (SV) replication in cultured Aedes albopictus (A. albo) or (A. aegypti (A. aeg) and BHK21 or chick embryo (CEF) cells. 1. Viral growth kinetics and yields are similar in A. albo cells at 28 degress C and in vertebrate cells at 37 degrees C. A. albo exhibit no CPE and yield persistenetly infected cultures. 2. SV grown in A. albo cells lacks sialic acid but is antigenically and in terms of particle/PFU or particle/HAU ratios equivalent to SV derived from vertebrate cells. The contrast to VSV in the latter respect is discussed. 3. SV from persistently infected A. albo or A. aeg cells is temperature-sensitive, thermolabile, and produces small plaques. Partial characterization of these mutants, of RNA associated with their replication, and their high reversion rate to ts+ upon serial undiluted passage in GHK21 cells are presented. 4. Hostdependent differences in the generation of defective-interfering (DI) SV particles and of low molecular weight viral RNA species have been observed upon undiluted serial passages in BHK21 and CEF. In contrast, serial passage in A. albo cells appears not to produce DI particles or small RNA species nor do these cells "recognize" as such DI particles from BHK21 cells. 5. Possible implications of these observations fro the natural life cycle of arthropod-borne togaviruses are discussed. [References: 23]

AU - Dubin DT, Stollar V
TI - Methylation of Sindbis virus "26S" messenger RNA.
SO - Biochem Biophys Res Commun 1975 Oct 27;66(4):1373-9

AU - Banerjee SN, Buchmeier M, Rawls WE
TI - Requirement of cell nucleus for the replication of an arenavirus.
SO - Intervirology 1975-76;6(3):190-6
AB - Baby hamster kidney (BHK21) cells enucleated with cytochalasin B were infected with Pichinde virus or Sindbis virus. Viral replication was measured by plaque assay, and the synthesis of viral antigens was determined by immunofluorescence. Pichinde virus replication was completely inhibited in cells enucleated prior to infection as measured by both techniques while the replication of Sindbis virus was unaffected. Enucleation of cells at different times after infection with Pichinde virus indicated that intact nuclei were required for at least 8 h after infection.

AU - Brawner TA, Steglich C, Sagik BP
TI - Genetic exclusion and stable complementation of Sindbis virus.
SO - Arch Virol 1976;50(3):177-87
AB - In an effort to enhance genetic interactions by eliminating spatial or physical barriers between variants of Sindbis virus MgCl2 was used to aggregate infecting viral particles. Mixing viral samples in a 1:1 ratio with 0.5 M MgCl2 produced maximal reduction in plaque forming units (PFU) with minimal cell damage due to MgCl2. Aggregate size was determined to be about 7 PFU. Samples taken at 3,5 and 10 hours after infection with mixed aggregates composed of large and small plaque forming virus indicated that only one type of genome was represented among the progeny particles. In addition, aggregation enhanced complementation and the progeny were stable after several cycles of sonication and passage.

AU - Igarashi A, Stollar V
TI - Failure of defective interfering particles of Sindbis virus produced in BHK or chicken cells to affect viral replication in Aedes albopictus cells.
SO - J Virol 1976 Aug;19(2):398-408
AB - Whereas defective interfering particles of Sindbis virus are readily produced in BHK-21 cells or chicken embryo fibroblasts by the techniques of serial undiluted passage, similar methods failed to generate such particles in Aedes albopictus cell cultures. In addition, Sindbis virus stocks produced in BHK-21 cells or chicken embryo fibroblasts and which contained defective interfering particles, when tested in A. albopictus cells, failed (i) to interfere with the replication of standard Sindbis virus and (ii) to change the pattern of intracellular viral RNA synthesis from that produced by infection with standard Sindbis virus alone. We conclude that defective interfering particles of Sindbis virus generated in chicken or hamster cells are silent or inert in mosquito cells.

AU - Margotat A, Laplane J, Pisano MR, Nicoli J
TI - [Polyuridylic sequences and negative strands of Sindbis virus-specific RNAs: study by affinity chromatography on poly (A)-sepharose columns (author's transl)]. [French]
SO - Ann Microbiol (Paris) 1976 Aug-Sep;127B(2):243-56
AB - The experimental conditions of affinity chromatography on poly (A)-Sepharose columns have been determined. This method makes obvious the existence of polyuridylic acid sequences on the negative strands of Sinbis virus-spedific RNAs. The isolated RNAs are negative and positive strands hybrids. By polyacrylamide gel electrophoresis, it has been shown that the negative strands have the same length as the 26 S interjacent RNA at the 6th hour, and as the 42 S virion RNA at the 9th hour postinfection. The polyadenylic acid sequences of the virion RNA and of the replicative intermediate are therefore probably genetically coded.

AU - Bracha M, Leone A, Schlesinger MJ
TI - Formation of a Sindbis virus nonstructural protein and its relation of 42S mRNA function.
SO - J Virol 1976 Dec;20(3):612-20
AB - Chicken embryo fibroblasts infected with an RNA- temperature-sensitive mutant (ts24) of Sindbis virus accumulated a large-molecular-weight protein (p200) when cells were shifted from the permissive to nonpermissive temperature. Appearance of p200 was accompanied by a decrease in the synthesis of viral structural proteins, but [35S]methionine tryptic peptides from p200 were different from those derived from a 140,000-molecular-weight polypeptide that contains the amino acid sequences of viral structural proteins. Among three other RNA- ts mutants that were tested for p200 formation, only one (ts21) produced this protein. The accumulation of p200 in ts24- and ts21-infected cells could be correlated with a shift in the formation of 42S and 26S viral RNA that led to an increase in the relative amounts of 42S RNA. These data indicate that p200 is translated from the nonstructural genes of the virion 42S RNA and further suggest that this RNA does not function effectively in vivo as an mRNA for the Sindbis virus structural proteins.

AU - Bracha M, Schlesinger MJ
TI - Inhibition of Sindbis virus replication by zinc ions.
SO - Virology 1976 Jul 1;72(1):272-7

AU - Atkins GJ
TI - The effect of infection with Sindbis virus and its temperature-sensitive mutants on cellular protein and DNA synthesis.
SO - Virology 1976 Jun;71(2):593-7

AU - Kimura Y, Norrby E, Nagata I, Ito Y, Shimokata K
TI - Homologous interference induced by a temperature-sensitive mutant derived from an HVJ (Sendai virus) carrier culture.
SO - J Gen Virol 1976 Nov;33(2):333-43
AB - Homologous interference between a temperature-sensitive small plaque mutant (HVJ-pB) derived from an HVJ (haemagglutinating virus of Japan - the Sendai strain of parainfluenza I virus) carrier culture of BHK cells and the original wild-type virus (HVJ-W) has been investigated. Prior infection of LLCMK2, HeLa, BHK or mouse L cells with HVJ-pB, both at permissive and non-permissive temperatures, for 24 h resulted in a reduced yield of superinfecting HVJ-W, reflecting a smaller number of cells capable of producing the superinfecting virus. However, HVJ-pB did not interfere with the replication of vesicular stomatitis virus, Sindbis virus or Newcastle disease virus. Interference in this system seems to be due to inhibition of the attachment of superinfecting HVJ-W as a result of intracellular mechanisms operating at a late stage in the replication of the interfering virus. There is also blocking or destruction of cellular receptors by extra-cellular particles of the interfering virus. Protein synthesis coded for by the complete virus genome is required to establish and maintain the interference, and treatment with actinomycin D has no effect on the interference phenomenon.

AU - Bras-Herreng F
TI - [Adaptation of a Sindbis virus population to "Drosophila melanogaster" (author's transl)]. [French]
SO - Ann Microbiol (Paris) 1976 Nov-Dec;127B(4):541-65
AB - The evolution of a Sindbis virus population during serial passages in drosophila has been studied. The adaptation of Sindbis virus to this unusual host was achieved within about twenty passages. Of course, the viral population went on evolving: a thermosensitive mutant appeared which excluded the original virus; later, a new type, different in plaques morphology, was observed: it did not exclude the former type but remained balanced with it. The adaptation of Sindbis virus to drosophila was revealed in two ways: first, in an improvement of the efficiency of infection in flies without any alteration of the plating efficiency on vertebrate cells; second, in an earlier and earlier invasion of flies which resulted from an accelerated rate of early virus production by the cells. A relationship between those two modifications is suggested. The adaptation of Sindbis virus to drosophila was probably the result of a rather high number of mutationnal steps which did not involve any concomitant change in the thermosensitivity of the viral population. An interpretation is presented.

AU - Dikii VV, Petkevich AS, Leont'eva NA, Galegov GA
TI - [Effect of rimantadine on the synthesis of virus-specific RNA in the culture of cells infected with Sindbis virus]. [RUSSIAN]
SO - Vopr Med Khim 1976 Nov-Dec;22(6):844-8
AB - Effect of rimanthadine (alpha-methyl-I-adamantane methylamine) on the synthesis of virus specific RNA was studied in culture of cells, infected with Syndbis virus. Rimanthadine inhibits the synthesis of virus specific RNA beginning from the 3-rd hr of infection. A magnitude of inhibition from the 3-rd to the 7-th hrs of infection was practically the same and exceeded 50%. In the presence of rimanthadine "early" RNA was formed (with sedimentation constant of 20-14 S), where radioactivity was lowered as compared with control; formation of more "late" RNA peaks (43 S, 34 S and 26 S) was completely prevented within these periods of infection.

AU - Zhdanov VM, Azadova NB
TI - [Integration and transfection of an arbovirus by mammalian cells]. [RUSSIAN]
SO - Mol Biol (Mosk) 1976 Nov-Dec;10(6):1296-302
AB - A system: L cells chronically infected with Sindbis virus was studied. Unlike acute infection wherein the mature virions are produced, the chronically infected tissue culture produces subviral structures-infectious ribonucleoproteins. Molecular hybridization experiments revealed the integration of the viral genome (DNA-transcript) into the cellular genome. Transfection experiments showed the possibility to induce the synthesis of the virus in sensitive cells treated with DNA from the chronically infected cells.

AU - Strauss EG, Lenches EM, Strauss JH
TI - Mutants of sindbis virus. I. Isolation and partial characterization of 89 new temperature-sensitive mutants.
SO - Virology 1976 Oct 1;74(1):154-68

AU - Renz D, Brown DT
TI - Characteristics of Sindbis virus temperature-sensitive mutants in cultured BHK-21 and Aedes albopictus (Mosquito) cells.
SO - J Virol 1976 Sep;19(3):775-81
AB - A number of the temperature-sensitive mutants of Sindbis virus originally isolated and characterized by Burge and Pfefferkorn (1966, 1968) were reexamined for their abilities to grow and complement one another in cultured BHK-21 and Aedes albopictus (mosquito) cells. The response of the mutants to conditions of high and low temperature was similar in cultured cells of both the vertebrate and invertebrate hosts. Complementation experiments in BHK-21 cells produced growth patterns similar to those described by Burge and Pfefferkorn for chicken embryo fibroblast cells (1966) and placed the mutants into six nonoverlapping complementation groups. When examined in the cultured mosquito cells, only three of the nine mutants used in this study demonstrated complementation under a variety of experimental conditions. Homologous interference experiments demonstrated that the unusual patterns of complementation obtained in the A. albopictus cells did not result from an inefficient infection of the invertebrate cells by the mutants.

AU - Pogodina VV, Kiseleva LL, Fokina GI, Korolev MB, Sensiuta NB
TI - [Interaction of the Sindbis virus and Rauscher and Friend leukoviruses in primary cultures and subcultures of mouse fibroblasts in the early stages of persistence]. [RUSSIAN]
SO - Vopr Virusol 1977;(1):44-50
AB - The interaction between Friend and Raucher leukoviruses and Sindbis togavirus was studied in primary cultures of mouse fibroblasts and subcultures passaged for 77 days. In primary cultures, two types of virus interactions were observed: neutralism and interference. In interference, the release of the infectious Sindbis virus from the cells is blocked. According to electron microscopic observations, its reproduction terminates by formation of virus nucleocapsid. The blocking of the togavirus maturation is stable in primary cultures but reversible upon subcultivation of the cells infected with oncorna- and togavirus. Rauscher and Sindbis viruses are capable of joint persistence in subcultures with a gradual decrease of the infectivity of togavirus and the leukemogenic activity of oncornavirus.

AU - Martire G, Bonatti S, ALIPERTI G, De Giuli C, Cancedda R
TI - Free and membrane-bound polyribosomes in BHK cells infected with Sindbis virus.
SO - J Virol 1977 Feb;21(2):610-8
AB - The data presented in the paper demonstrate that in BHK cells infected with Sindbis virus virtually all the 42S mRNA not in nucleocapsid is associated with free polyribosomes, whereas the 26S mRNA is distributed between free and membrane-bound polyribosomes. We suggest that the 26S RNA polyribosomes are bound to the membranes through the nascent chains of the B1 protein and that a large percentage of 26S RNA polyribosomes free in the cytoplasm may be due to the small amount of rough endoplasmic reticulum in BHK cells. In addition, we found that intracellular nucleocapsid is in the nonmembrane fraction of the cytoplasm of infected cells.

AU - Schluter B, Vosdingh R, Brown A
TI - Pathogenesis of temperature-sensitive mutants of Sindbis virus in the embryonated egg. III. Autologous, homologous, and heterologous interference.
SO - J Infect Dis 1977 Jul;136(1):7-16
AB - Two temperature-sensitive (ts) mutants of Sindbis virus able to synthetize RNA, but not two ts mutants defective in viral RNA synthesis, reduced to lethality of the superinfecting heat-resistant parent strain of Sindbis virus in embryonated chicken eggs as compared with results in controls. Autologous interference (of mutant with parent) was observed under a variety of conditions. Time-course studies in one selected system showed that lower titers of superinfecting parent virus were detected in doubly infected eggs than in controls eggs infected with only parent virus. Homologous interference with lethality (of mutant with an immunologically different yet related group A togavirus) was observed when the interfering virus was a ts mutant of either Sindbis virus or Eastern equine encephalitis virus. Heterologous interference (mutant with unrelated virus) could also be demonstrated with a ts mutant of Sindbis virus against vaccinia virus-induced pock formation or death. In an autologous interference experiment, in which the ts mutant of Sindbis virus was administered by the yolk sac route and the egg was challeged with parent heat-resistant Sindbis virus by the allantoic route, an interferon-like substance was found in the allantoic fluids at the time of challenge. It is concluded that certain ts viruses of togavirus group A can induce autologous, homologous, and heterologous viral interference in vivo.

AU - Sarver N, Stollar V
TI - Sindbis virus-induced cytopathic effect in clones of Aedes albopictus (Singh) cells.
SO - Virology 1977 Jul 15;80(2):390-400

AU - Ortin J, Vinuela E
TI - Requirement of cell nucleus for African swine fever virus replication in Vero cells.
SO - J Virol 1977 Mar;21(3):902-5
AB - The role of the cell nucleus in the development of African swine fever virus in Vero cells has been studied. No viral growth could be detected in enucleated cells under conditions that allow normal development of Sindbis virus. Furthermore, African swine fever virus DNA synthesis was inhibited more than 95% after infection of enucleated Vero cells as compared with normal cells.

AU - Brzeski H, Kennedy SI
TI - Synthesis of Sindbis virus nonstructural polypeptides in chicken embryo fibroblasts.
SO - J Virol 1977 May;22(2):420-9
AB - The identification of eight previously undescribed polypeptides in chicken embryo cells infected with Sindbis virus is reported. Seven of these polypeptides were distinguishable from the virus structural polypeptides and their precursors by their molecular weights and tryptic peptide maps. The eighth was closely related to pE2 (Schlesinger and Schlesinger, 1973), a precursor to one of the virus particle glycoproteins. Pulse-chase experiments and the use of an inhibitor of proteolytic cleavage allowed a division of the seven nonstructural (NS) polypeptides into three stable end products (NS p89, NS p82, and NS p60) and four precursors (p230, p215, p150, and p76). The labeling kinetics after synchronous initiation of translation indicated that synthesis of the NS polypeptides started at a single site and showed that the order of the genes coding for the NS polypeptides was (5' leads to 3') NS p60, NS p89, and NS p82. Short-pulse experiments under conditions of both synchronized and nonsynchronized translation suggested that cleavage of the primary translation product of the NS genes occurred only after its synthesis was completed and that the first cleavage removed the C-terminal polypeptide. From these and other experiments, we propose a detailed scheme for the synthesis and processing of Sindbis virus NS polypeptides.

AU - Gushchin BV, Berezina LK, Gushchina EA, L'vov DK, Klimenko SM
TI - [Interaction of Sindbis virus with a culture of cells producing oncornavirus]. [RUSSIAN]
SO - Vopr Virusol 1977 May-Jun;(3):326-31
AB - Electron microscopy and biophysical methods were used for examinations of primarily trypsinized chick fibroblast cell culture spontaneously producing C-type oncornavirus at 1, 6, and 24 hours after inoculation with Sindbis virus. During the first 6 hours rapid maturation and release of oncornavirus from cells were observed. At later intervals oncornavirus production was inhibited. It is assumed that in the system under study, biosynthesis of oncornavirus and Sindbis virus occur separately.

AU - Schlesinger RW, Stollar V, Guild GM, Igarashi A, Shenk TE, Peleg J
TI - The significance and nature of defective interfering viruses.
SO - Bull Schweiz Akad Med Wiss 1977 Sep;33(4-6):229-42
AB - Deletions in viral genomes appear to be a common occurrence in the replication of all DNA and RNA viruses which have been adequately studied. Such defective genomes can replicate in the presence in the same cell of a helper virus as long as the deletion does not involve the initiation site for genome replication. Coinfection of a cell with defective and "normal" infectious virus leads to reduction in the yield of the latter. The nature of DI viruses and genomes found in Sindbis virus-infected vertebrate cells during "undiluted passage" series is discussed. This procedure leads to the accumulation of progressively shorter viral RNA genomes with internal deletions. The enrichment is limited to genome lengths which are integral fractions (1/2, 1/3, 1/4, etc.) of the complete genome, and these are also found in viral particles released at the corresponding passage levels. It is believed that the selective accumulation of these fragments is governed by constraints of assembly which demand that one full genome equivalent be packaged in a released particle. In contrast to vertebrate cells, cultured mosquito cells do not seem to produce or "recognize" DI particles. Possible implications for the epidemiology of arthropod-transmitted alphaviruses are presented.

AU - Stollar V
TI - Inhibition of Sindbis virus replication in Aedes albopictus cells deprived of methionine.
SO - Virology 1978 Dec;91(2):504-7

AU - Sarver N, Stollar V
TI - Virazole prevents production of Sindbis virus and virus-induced cytopathic effect in Aedes albopictus cells.
SO - Virology 1978 Dec;91(2):267-82

AU - Peleg J, Pecht M
TI - Adaptation of an Aedes aegypti mosquito cell line to growth at 15 degrees C and its response to infection by Sindbis virus.
SO - J Gen Virol 1978 Feb;38(2):231-9
AB - Aedes aegypti mosquito cells, usually cultured at 28 to 30 degrees C, were adapted to grow at 15 degrees C. They were designated A. aegypti (c) cells, and had an estimated doubling time of 10 days. Sindbis virus (SV) replicated in these cells to peak titres of over 1.0 x 10(9) p.f.u./ml 8 to 10 days after inoculation. These, or about 10-fold lower titres, continued to be produced over a 130 day test period without causing visible cell damage. Continuous virus proliferation and the yield of uniformly large plaque forming progeny viruses are the two most important features which differentiate infection with this virus in A. aegypti (c) cells from that of A. aegypti cells grown at 28 degrees C (Peleg & Stollar, 1974). Absence of homologous interference vis-a-vis cell-virus coexistence suggests that homologous interference is not a prerequisite for maintaining cell-virus coexistence. Preinoculation of A. aegypti (c) cultures with a small plaque forming Sindbis virus (SV-S) leads, under certain conditions, to the establishment of homologous interference.

AU - Pogodina VV, Kiseleva LL, Miller GG, Fokina GI, Graevskaia NA
TI - [Persistence of Sindbis virus in cultures either non-producing or irregularly producing oncornavirus]. [RUSSIAN]
SO - Vopr Virusol 1978 Jan-Feb;(1):52-6
AB - Persistence of Sindbis virus (SV) was studied for 9 months in two lines of mouse cell cultures (BALB/C) in one of which the genome of endogenous ecotropic oncornaviruses was repressed. The other lines was exogenously infected at the level of plimary culture with Rauscher leukemia virus (RLV) and SV and showed gradual inhibition of oncornavirus functions. The presence of oncornavirus type C was not the necessary condition for the development of persistent SV infection, however it influenced the character of persistence. In both systems, sequential loss of the hemagglutinating and interferon-inducing activities, then infectivity of SV (61--82 days), and persistence of the noninfectious antigen of the arbovirus for 9 months were observed. The differences consisted in the time of appearance of homologous interference to SV: in the presence of oncornavirus earlier (40 days), under conditions of repressed oncornavirus genome later (179 days). Electron microscopic examinations showed that in the system infected with RLV and SV there occurred in the course of persistence a sharp activation of phagosome-lysosome complex accompanied by incorporation into phagocytolysomes of numerous intact and partially destroyed virions of SV and RLV and their release from cell with cytoplasmic fragments. Possible mechanisms of inhibition of functions of the oncogenic and infectious viruses in the reported model of mixed chronic infection are discussed.

AU - Mento SJ, Stollar V
TI - Effect of ouabain on sindbis virus replication in ouabain-sensitive and ouabain-resistant Aedes albopictus cells (Singh).
SO - Virology 1978 Jun 1;87(1):58-65

AU - Brzeski H, Clegg JC, Atkins GJ, Kennedy SI
TI - Regulation of the synthesis of Sindbis virus-specified RNA: role of the virion core protein.
SO - J Gen Virol 1978 Mar;38(3):461-70
AB - Cells infected with seven different RNA+ mutants of Sindbis virus were found to accumulate a virus-specified polypeptide of mol. wt. 144000 (p144) during incubation at the non-permissive temperature, while at the same time synthesis of the virus structural proteins was drastically reduced. Mapping of the tryptic peptides of p144 showed that it contained the amino acid sequences of all the virus structural proteins. At the non-permissive temperature cells infected with the same seven mutants (out of 28 examined) also showed increased synthesis of 26S RNA, the mRNA for the virus structural proteins, relative to 42S RNA, and the virus genome, compared with infections by wild-type virus. We propose that both these phenotypic effects are the results of a single mutational step and that the primary defect in the processing of the virus structural protein precursor induces the relatively increased rate of synthesis of structural protein mRNA. Temperature-shift experiments with mutant-infected cells showed that p144 itself is not the agent of this effect. The failure of exposure to zinc ions to alter the RNA ratio in wild-type virus-infected cells suggested that the virus envelope proteins are not involved either, since their synthesis is preferentially inhibited under these circumstances. It is possible that it is the failure to synthesize the proper quantity of core protein in the mutant-infected cells which causes the shift of RNA synthesis in favour of structural protein mRNA.

AU - Pogodina VV, Fokina GI, Kiseleva LL, Graevskaia NA, Sito AF
TI - [Persistence of the Sindbis virus in cultures producing oncornavirus]. [RUSSIAN]
SO - Vopr Virusol 1978 Mar-Apr;(2):196-201
AB - Continuous lines were obtained from primary cultures of BALB/C mouse embryo cells which were found by electron microscope and reverse transcriptase reaction to produce permanently oncoronavirus type C after exogenous infection with Rauscher leukemia virus (RLV). Sindbis virus (SV) was inoculated into virogenic cultures 398 days after infection with RLV. The system in characterized by rapid (3-21 days) disappearance of the infectious arbovirus from the medium and the cells, long-term (over 5 months) persistence on SV noninfectious antigen and signs of stimulation of oncornavirus activity. The level of reverse transcriptase activity in cultures in the presence of persisting arbovirus was 1.5-3.3-fold higher than in cultures infected with RLV alone. Two variants of the course of mixed chronic infection of the cultures with oncornavirus and arbovirus differing in the rate of transition of the arbovirus into the noninfectious form and inhibition or stimulation of oncornavirus functions are discussed.

AU - Igarashi A
TI - Isolation of a Singh's Aedes albopictus cell clone sensitive to Dengue and Chikungunya viruses.
SO - J Gen Virol 1978 Sep;40(3):531-44
AB - Twenty clones were isolated from cultured Aedes albopictus (Singh) cells in the presence of anti-Chikungunya (CHIK) virus serum. Each clone was tested for its yields of Dengue (DEN) viruses, types 1, 2, 3 and 4, and also CHIK virus. Clone C6 showed the highest yield of each virus tested. Forty-three clones obtained by recloning C6 in the presence of anti-DEN sera showed almost the same virus yields as C6. One of the clones, C6/36, showed mild to extensive cytopathic effects several days after virus infection, in contrast to the original uncloned (SAAR) cells. Fluorescent antibody staining revealed that the amount of virus antigen accumulated in the cytoplasm was almost the same in every cell in the case of clone C6/36, while it was highly heterogeneous for uncloned SAAR cells. Growth curves of the viruses indicated that clone C6/36 gave a significantly higher yield for each virus than uncloned SAAR cells up to 7 days after infection. Virus sensitivity of the C6/36 clone did not change by growing the cells with the medium used for uncloned SAAR cells, nor did the virus sensitivity of uncloned cells increase in medium used for clone C6/36. However, the C6/36 clone became resistant to CHIK virus, but not to DEN or Sindbis viruses, after incubation with the medium used for another A. albopictus cell line (SAAK). The transfer of the specific resistance to CHIK may be mediated by some latent virus related to CHIK.

AU - Frey TK, Gard DL, Strauss JH
TI - Replication of Sindbis virus. VII. Location of 5-methyl cytidine residues in virus-specific RNA.
SO - Virology 1978 Sep;89(2):450-60

AU - Martin JH, Weir RC, Dalgarno L
TI - Replication of standard and defective Ross River virus in BHK cells: patterns of viral RNA and polypeptide synthesis.
SO - Arch Virol 1979;61(1-2):87-103
AB - Virus-specific macromolecule synthesis has been examined in BHK cells infected with Ross River virus. Unpassaged virus (R-0) and tenth-passage virus (R-10) have been compared. In infected cells R-0 generates i) 45S, 28S, 33S and 26S viral RNAs, ii) virus-specific precursor polypeptides of mol. wt. 127,000, 95,000 and 61,000 and iii) viral envelope proteins (mol. wts. 52,000 and 49,000) and nucleocapsid protein (mol. wt. 32,000). Thus in terms of virus-specific RNA and polypeptide synthesis, the replication of standard RRV is analogous to that of Semliki Forest virus and Sindbis virus. R-10 interferes with the replication of standard Ross River virus and generates large amounts of 19S and 24S defective RNA species; 45S and 26S RNA synthesis was not markedly affected. Defective RNAs are associated with RNAse-sensitive, 50S cytoplasmic particles which contain a variety of (mainly host) proteins but no nucleocapsid protein. No evidence for translation of defective RNAs was obtained. R-10 infection is also characterized by a relatively early shut down of host protein syntehsis and by a reduction in virus-specific polypeptide synthesis and nucleocapsid formation. The data suggest that defective Ross River virus interferes primarily at the translational level.

AU - Martin JD, Riggsby WS, Beck RW
TI - The effect of ribonuclease on the replicative forms of Sindbis virus RNA.
SO - Arch Virol 1979;60(2):131-46
AB - Three species of double-stranded RNA, designated RF I, RF II, and RF III in order of decreasing size (25), are produced by ribonuclease treatment of extracts of chicken embryo cells infected for 6 hours with Sindbis virus. Only one class of replicative form RNA is present in extracts not treated with ribonuclease; this class contains some molecules which can be enzymatically cleaved to produce the other two replicative forms. At a low level of enzyme (0.001 microgram/ml) the major species obtained was RF I, the replicative form of the genome. When the enzyme concentration was increased 10-, 100-, and 1000-fold, there was a progressive increase in the proportions of RF's II and III and a concomitant decrease in the proportion of RF I. The generation of RF's II and III by nuclease resulted in the ratio expected for these two species if they are produced by cleavage of RF I-like molecules. In preparations of isolated double-stranded RNA, only RF I and replicative intermediate RNA were present. Mild nuclease treatment of these preparations converted the replicative intermediates primarily to RF I. Higher enzyme levels generated greater proportions of RF II and RF III, but RF I-like molecules were the major source for these increased proportions. Treatment of the isolated naturally occurring replicative form with 0.01 microgram of ribonuclease per ml cleaved some molecules migrating as RF I during gel electrophoresis into molecules which migrated as RF II and RF III.

AU - Eaton BT
TI - Heterologous interference in Aedes albopictus cells infected with alphaviruses.
SO - J Virol 1979 Apr;30(1):45-55
AB - Maximum amounts of 42S and 26S single-stranded viral RNA and viral structural proteins were synthesized in Aedes albopictus cells at 24 h after Sindbis virus infection. Thereafter, viral RNA and protein syntheses were inhibited. By 3 days postinfection, only small quantities of 42S RNA and no detectable 26S RNA or structural proteins were synthesized in infected cells. Superinfection of A. albopictus cells 3 days after Sindbis virus infection with Sindbis, Semliki Forest, Una, or Chikungunya alphavirus did not lead to the synthesis of intracellular 26S viral RNA. In contrast, infection with snowshoe hare virus, a bunyavirus, induced the synthesis of snowshoe hare virus RNA in both A. Ablpictus cells 3 days after Sindbis virus infection and previously uninfected mosquito cells. These results suggested that at 3 days after infection with Sindbis virus, mosquito cells restricted the replication of both homologous and heterologous alphaviruses but remained susceptible to infection with a bunyavirus. In superinfection experiments the the alphaviruses were differentiated on the basis of plaque morphology and the electrophoretic mobility of their intracellular 26S viral RNA species. Thus, it was shown that within 1 h after infection with eigher Sindbis or Chikungunya virus, A. albopictus cells were resistant to superinfection with Sindbis, Chikungunya, Una, and Semliki Forest viruses. Infected cultures were resistant to superinfection with the homologous virus indefinitely, but maximum resistance to superinfection with heterologous alphaviruses lasted for approximately 8 days. After that time, infected cultures supported the replication of heterologous alphaviruses to the same extent as did persistently infected cultures established months previously. However, the titer of heterologous alphavirus produced after superinfection of persistently infected cultures was 10- to 50-fold less than that produced by an equal number of previously uninfected A. albopictus cells. Only a small proportion (8 to 10%) of the cells in a persistently infected culture was capable of supporting the replication of a heterologous alphavirus.

AU - Ng ML, Westaway EG
TI - Proteins specified by togaviruses in infected Aedes albopictus (Singh) mosquito cells.
SO - J Gen Virol 1979 Apr;43(1):91-101
AB - Yields of greater than 10(7) p.f.u./ml at 28 or 37 degrees C of the alphavirus Sindbis and the flavivirus Kunjin were obtained in the Aedes Albopictus (Singh) cell line, the latent periods being 4 to 6 and 10 to 12 h, respectively. Despite a high background of host protein synthesis, virtually all the virus-specified proteins of the flaviviruses Kunjin, Dengue-2 and Japanese encephalitis were labelled and resolved by slab gel electrophoresis of infected and uninfected cell proteins. In contrast, only one induced protein, of mol. wt. 30 000, was identified in cells labelled during Sindbis virus infection. The envelope glycoprotein V3 of Kunjin virus was resolved as a double band in samples of infected cytoplasm labelled with 3H-glucosamine, similar to that of carbohydrate-labelled V3 in vertebrate (Vero) cells. Attempts to reduce host protein synthesis selectively during labelling periods were unsuccessful using either a hypertonic inhibition block or treatment with 0.1 mu g actinomycin D per ml. The most efficient labelling of Kunjin virus-specified proteins was achieved at 37 degrees C in the presence of actinomycin D. The largest non-structural flavivirus protein NV5 migrated slightly faster than NV5 from infected vertebrate (Vero) cells. The small non-structural proteins NV1, NV1 1/2 and NV2 from infected mosquito cells were successively trimmed during post translational periods exceeding 70 min, compared to much shorter periods reported previously for post translational modifications of these proteins in vertebrate cells.

AU - Barrett PN, Atkins GJ
TI - Virulence of temperature-sensitive mutants of Sindbis virus in neonatal mice.
SO - Infect Immun 1979 Dec;26(3):848-52
AB - The virulence in neonatal mice of temperature-sensitive (ts) mutants of Sindbis virus was determined by measurements of mean survival time and 50% lethal dose after intracerebral injection. For 11 ts mutants, mean survival time was determined by the ribonucleic acid (RNA) phenotype, RNA+ mutants killing the mice sooner than RNA- mutants for the same titer of virus injected. Mortality caused by seven ts mutants was, with one exception, correlated with the proportion of revertants recovered after death. A82, a presumed double mutant showing low reversion, showed no detectable lethality. The pathogenicity of this mutant could be detected by inhibition of weight gain, which was proportional to the titer of virus injected. A low-level persistence, independent of the titer injected, occurred up to 7 days after injection. This was followed by complete clearance. It is concluded that the virulence of Sindbis virus may be considerably altered by mutation, and that this is related to events occurring at the cellular level.

AU - Lombard Y, Poindron P, Porte A
TI - [Origin and formation of different types of vacuoles induced by the multiplication of the alphavirus Sindbis virus in various cell systems]. [French]
SO - Can J Microbiol 1979 Dec;25(12):1452-9
AB - Spherule-containing vacuoles and nucleocapsid-bearing vacuoles (cytopathic vacuoles types 1 and 2 respectively of Grimley et al. 1968) induced by Alphavirus Sindbis were studied in brains from newborn mice, chicken embryo fibroblasts, and two lines of tumoral glial cells from muridae. Endoplasmic reticulum (ER) elements and finely granular electron-dense material also seen in contact with nucleocapsids seemed to be involved in the formation of the classical single-membrane spherule-containing vacuoles. A second type of spherule-containing vacuoles were characterized by their double membrane and an amorphous electron-dense content and were probably derived from mitochondria. Nucleocapsid-bearing vacuoles were formed from modified ER elements and seemed to be linked to excessive synthesis of viral material. Such ER alterations were not observed in RG6 cells. In these cells, there were only spherule-containing vacuoles, while nucleocapsids were seen associated with the cytoplasmic membrane only.

AU - Cancedda R, Shatkin AJ
TI - Ribosome-protected fragments from sindbis 42-S and 26-S RNAs.
SO - Eur J Biochem 1979 Feb 15;94(1):41-50
AB - Sindbis virus 42-S and 26-S RNAs labeled with 32P were purified from infected chick embryo fibroblasts. The RNA's were incubated in the presence of a wheat germ cell-free translating system under conditions that yielded 40-S and 80-S initiation complexes. After digestion with RNase A, ribosome-protected fragments were isolated by polyacrylamide gel electrophoresis and compared with respect to number, size, cap content and oligonucleotide composition. The two RNA species yielded several fragments of chain length about 35--40 nucleotides from 80S complexes and up to 60--65 nucleotides from 40-S complexes. The 5'-terminal capped sequence, m7 GpppA-U-G that is present in both Sindbis virus RNA's, was not retained in any of the ribosome-protected fragments. Fingerprint analyses indicated that the fragments derived from 40S and 80-S initiation complexes of each species of RNA were overlapping, but the fragments from 42-S and 26-S RNAs were unrelated. The complexity of the fingerprints were consistent with protection of a single, different initiation site in each Sindbis virus RNA.

AU - Riedel B, Brown DT
TI - Novel antiviral activity found in the media of Sindbis virus-persistently infected mosquito (Aedes albopictus) cell cultures.
SO - J Virol 1979 Jan;29(1):51-60
AB - Aedes albopictus (mosquito) cells persistently infected with Sindbis virus for a period of 6 months release into the medium a low-molecular-weight material capable of specifically reducing the yields of Sindbis virus during the "acute phase" of infection in mosquito cells. The antiviral activity was produced in detectable levels at 3 days after infection, and its concentration in the extracellular medium increased thereafter. The antiviral activity was inactivated by treatment with the enzyme protease K and heat. It was not activated by treatment with antibody prepared against extracts of Sindbis virus-infected BHK-21 cells. The antiviral activity differs from interferon produced by vertebrate cells in that it is virus specific as well as cell specific.

AU - Garry RF, Westbrook K, Waite MR
TI - Differential effects of ouabain on host- and sindbis virus-specified protein synthesis.
SO - Virology 1979 Nov;99(1):179-82

AU - Atkins GJ
TI - Establishment of persistent infection in BHK-21 cells by temperature-sensitive mutants of Sindbis virus.
SO - J Gen Virol 1979 Oct;45(1):201-7
AB - Twelve temperature-sensitive (ts) mutants of Sindbis were examined for their ability to establish persistent infection in BHK-21 cells at 39 degrees C. Five of these mutants were able to initiate colony formation in infected cultures, which followed an extensive c.p.e. Two of the mutants were able to establish persistent infections which survived beyond the fifth cell passage p.i. The ability to initiate colony formation was correlated with low reversion of the ts mutation, or with ability to interfer with the multiplication of the wild-type virus. Virus released from persistently infected cultures was not temperature-sensitive. The restriction of virus multiplication in persistently infected cells operated prior to virus-specified RNA synthesis. It is concluded that in this system establishment of persistent infection depends on an inhibition of virus multiplication early in infection and occurs in only a small proportion of infected cells.

AU - Keranen S, Kaariainen L
TI - Functional defects of RNA-negative temperature-sensitive mutants of Sindbis and Semliki Forest viruses.
SO - J Virol 1979 Oct;32(1):19-29
AB - Defects in RNA and protein synthesis of seven Sindbis virus and seven Semliki Forest virus RNA-negative, temperature-sensitive mutants were studied after shift to the restrictive temperature (39 degrees C) in the middle of the growth cycle. Only one of the mutants, Ts-6 of Sindbis virus, a representative of complementation group F, was clearly unable to continue RNA synthesis at 39 degrees C, apparently due to temperature-sensitive polymerase. The defect was reversible and affected the synthesis of both 42S and 26S RNA equally, suggesting that the same polymerase component(s) is required for the synthesis of both RNA species. One of the three Sindbis virus mutants of complementation group A, Ts-4, and one RNA +/- mutant of Semliki Forest virus, ts-10, showed a polymerase defect even at the permissive temperature. Seven of the 14 RNA-negative mutants showed a preferential reduction in 26S RNA synthesis. The 26S RNA-defective mutants of Sindbis virus were from two different complementation groups, A and G, indicating that functions of two viral nonstructural proteins ("A" and "G") are required in the regulation of the synthesis of 26S RNA. Since the synthesis of 42S RNA continued, these functions of proteins A and G are not needed for the polymerization of RNA late in infection. The RNA-negative phenotype of 26S RNA-deficient mutants implies that proteins regulating the synthesis of this subgenomic RNA must have another function vital for RNA synthesis early in infection or in the assembly of functional polymerase. Several of the mutants having a specific defect in the synthesis of 26S RNA showed an accumulation of a large nonstructural precursor protein with a molecular weight of about 200,000. One even larger protein was demonstrated in both Semliki Forest virus- and Sindbis virus-infected cells which probably represents the entire nonstructural polyprotein.

AU - Dubin DT, Timko K, Gillies S, Stollar V
TI - The extreme 5'-terminal sequences of sindbis virus 26 and 42 S RNA.
SO - Virology 1979 Oct 15;98(1):131-41

AU - Hirsch RL, Griffin DE
TI - The pathogenesis of Sindbis virus infection in athymic nude mice.
SO - J Immunol 1979 Sep;123(3):1215-8

AU - Sonenberg N, Rupprecht KM, Hecht SM, Shatkin AJ
TI - Eukaryotic mRNA cap binding protein: purification by affinity chromatography on sepharose-coupled m7GDP.
SO - Proc Natl Acad Sci U S A 1979 Sep;76(9):4345-9
AB - A 24,000-dalton polypeptide that binds strongly and can be specifically crosslinked to the 5'-terminal cap structure m7GpppN in eukaryotic mRNAs has been detected in protein synthesis initiation factor preparations [Proc. Natl. Acad. Sci. USA (1978) 75, 4843--4847]. This polypeptide has been purified to apparent homogeneity by one chromatographic passage through an affinity resin prepared by coupling the levulinic acid O2',3'-acetal of m7GDP to AH-Sepharose 4B. Translation, in HeLa cell extracts, of capped mRNAs including Sindbis virus, reovirus, and rabbit globin mRNAs was stimulated by the cap-binding protein under conditions that did not increase translation of noncapped RNAs of encephalomyocarditis virus and satellite tobacco necrosis virus.

AU - Czarniecki CW, Sreevalsan T
TI - Sindbis virus RNA replication. I. Properties of the 38s RNA species.
SO - J Gen Virol 1979 Sep;44(3):759-71
AB - Four species of single-stranded virus RNA (49S, 38S, 33S and 26S) were detected in chick embryo fibroblasts infected with Sindbis virus. The relative amounts of these RNAs were unaffected by the m.o.i. There was also no significant difference in the molar proportions of the four RNA species when purified virion RNA was used as the inoculum. These findings suggest that the 38S and 33S species represent products of the transcription of non-defective virion RNAs. Kinetic analyses of RNA synthesis indicated that during a 1 min pulse more radioactivity was associated with the 38S than with the 49S RNA and as the length of the pulse increased, the ratio of 38S/49S decreased, with the 49S appearing as the predominant species. Furthermore, addition of cycloheximide within the first 3 h p.i. resulted in detection of only the 49S species. Synthese of all four species was unaffected when the drug was added after this time period. These data suggest that the 38S species may represent newly synthesized 49S molecules and some protein(s) synthesized within the first 3 h p.i. is necessary for maintaining the 38S conformational form.

AU - Malinoski F, Stollar V
TI - Inhibition of Sindbis virus replication in Aedes albopictus cells by virazole (ribavirin) and its reversal by actinomycin: a correction.
SO - Virology 1980 Apr 30;102(2):473-6

AU - Fuller FJ, Marcus PI
TI - Sindbis virus. I. Gene order of translation in vivo.
SO - Virology 1980 Dec;107(2):441-51

AU - Bonatti S, Sonenberg N, Shatkin AJ, Cancedda R
TI - Restricted initiation of protein synthesis on the potentially polycistronic Sindbis virus 42 S RNA.
SO - J Biol Chem 1980 Dec 10;255(23):11473-7
AB - Sindbis 42 S genome RNA was isolated from virions and translated in vitro before and after purification by oligo(dT)-cellulose chromatography and sucrose density gradient centrifugation under denaturing conditions. In intact 42 S RNA, only the 5'-proximal initiation site for the synthesis of nonstructural proteins was used. The internally located start site for viral structural protein formation was active in broken genome RNA molecules where, as a consequence of fragmentation, it was closer to a 5' end. The results of several kinds of experiments indicate that the fragmentation-dependent synthesis of structural proteins directed by virion RNA was not due to the presence of 26 S subgenomic messenger RNA.

AU - Czarniecki CW, Sreevalsan T
TI - Sindbis virus RNA replication. II. Strand composition and metabolic fate of the multi-stranded RNA species.
SO - J Gen Virol 1980 May;48(1):75-85
AB - Double-stranded RNA from SB virus-infected cells was denatured and analysed on agarose-methylmercuric hydroxide gels. Equimolar amounts of three single-stranded species with mol. wt. of 4 x 10(6), 2.5 x 10(6) and 1.8 x 10(6) were found. Pulse and chase experiments in infected cells established a precursor-product relationship between the multi-stranded and single-stranded virus RNA species. The present results support the model in which the 49S and 26S species of virus RNA are synthesized in infected cells from two distinct replicating structures.

AU - Lazymova ZA, Linitskaia GL, Leont'eva NA, Galegov GA
TI - [Action of remantadine on virus-specific polypeptide synthesis in a Sindbis virus-infected cell culture]. [RUSSIAN]
SO - Vopr Med Khim 1980 Sep-Oct;26(5):677-9
AB - Effect of remantadine (alpha-methyl-I-adamantane methylamine) on reproduction and synthesis of the virus-specific polypeptides was studied in culture of chicken embryo fibroblasts, infected with virus Syndbis. The preparation inhibited the virus reproduction as well as the formation of virus-specific proteins after addition into the cultural medium together with virus, immediately after absorption and within 1 and 2 hrs after absorption. The data obtained suggest a possibility of an immediate effect of remantadine on the synthesis of virus-specific macromolecules.

AU - Galegov GA, Lazymova ZA, Linitskaia GL, Leont'eva NA
TI - [Combined effect of remantadine (alpha-methyl-1-adamantane methylamine) and ribovirin (1-beta-D-ribofuranosyl-1,2,4-triazole-3-carboxamide) on the reproduction of Sindbis virus in cell culture]. [RUSSIAN]
SO - Vopr Virusol 1980 Sep-Oct;(5):608-11
AB - The influence of remantadine, ribovirine, and combination thereof on Sindbis virus reproduction in cell cultures was studied. Comparison of remantadine and ribovirine showed the former to be a stronger inhibitor of Sindbis virus reproduction. The results of the combined effect of both drugs in chick embryo fibroblast cultures showed undoubtful advantages of this treatment over the use of each of the drugs alone. In studies of the effect of ribovirine on synthesis of virus-specific proteins and RNA no inhibition of formation of virus-specific macromolecules was observed.

AU - Baric RS, Trent DW, Johnston RE
TI - A Sindbis virus variant with a cell-determined latent period.
SO - Virology 1981 Apr 15;110(1):237-42

AU - Malinoski F, Stollar V
TI - Inhibitors of IMP dehydrogenase prevent sindbis virus replication and reduce GTP levels in Aedes albopictus cells.
SO - Virology 1981 Apr 30;110(2):281-9

AU - Scheefers-Borchel U, Scheefers H, Edwards J, Brown DT
TI - Sindbis virus maturation in cultured mosquito cells is sensitive to actinomycin D.
SO - Virology 1981 Apr 30;110(2):292-301

AU - Sawicki DL, Sawicki SG, Keranen S, Kaariainen L
TI - Specific Sindbis virus-coded function for minus-strand RNA synthesis.
SO - J Virol 1981 Aug;39(2):348-58
AB - The synthesis of minus-strand RNA was studied in cell cultures infected with the heat-resistant strain of Sindbis virus and with temperature-sensitive (ts) belonging to complementation groups A, B, F, and G, all of which exhibited an RNA-negative (RNA-) phenotype when infection was initiated and maintained at 39 degrees C, the nonpermissive temperature. When infected cultures were shifted from 28 degrees C (the permissive temperature) to 39 degrees C at 3 h postinfection, the synthesis of viral minus-strand RNA ceased in cultures infected with ts mutants of complementation groups B and F, but continued in cultures infected with the parental virus and mutans of complementation groups A and G. In cultures infected with ts11 of complementation group B, the synthesis of viral minus-strand RNA ceased, whereas the synthesis of 42S and 26S plus-strand RNAs continued for at least 5 h after the shift to 39 degrees C. However, when ts11-infected cultures were returned to 28 degrees C 1 h after the shift to 39 degrees C, the synthesis of viral minus-strand RNA resumed, and the rate of viral RNA synthesis increased. The recovery of minus-strand synthesis translation of new proteins. We conclude that at least one viral function is required for alphavirus minus-strand synthesis that is not required for plus-strand synthesis. In cultures infected with ts6 of complementation group F, the syntheses of both viral plus-strand and minus-strand RNAs were drastically reduced after the shift to 39 degrees C. Since ts6 failed to synthesize both plus-strand and minus-strand RNAs after the shift to 39 degrees C, at least one common viral component appears to be required for the synthesis of both minus-strand and plus-strand RNAs.

AU - Eaton BT
TI - Viral interference and persistence in Sindbis virus infected Aedes albopictus cells. [Review]
SO - Can J Microbiol 1981 Jun;27(6):563-7

AU - Ivannikova TA, Karmysheva VYa, Ovsyannikova NV, Farutina LM
TI - Chronic alphavirus infection of L cells.
SO - Acta Virol 1981 Mar;25(2):95-100
AB - Chronic infection of L cells with Sindbis virus, induced in the absence of antiviral antibody, was studied by virological methods in combination with light and electron microscopy. Both Sindbis virus and endogenous oncovirus antigens were revealed in the cells by immunofluorescence. Sindbis virus and interferon were detected in the culture fluid. Immunoelectron microscopy revealed aggregation of Sindbis virus particles by immune serum.

AU - Barrett PN, Atkins GJ
TI - Establishment of persistent infection in mouse cells by Sindbis virus and its temperature-sensitive mutants.
SO - J Gen Virol 1981 May;54(Pt 1):57-65
AB - The ability of wild-type (wt) Sindbis virus and six temperature-sensitive (ts) mutants to establish persistent infection in mouse L cells and a line of mouse embryo (ME) cells was determined. The wt established persistent infection in both ME cells and L cells at 39 degrees C. At 30 degrees C the wt established persistent infection in L cells but not ME cells, which did not recover from the initial infection. For the ts mutants, both cell lines survived the initial infection at 39 degrees C (the restrictive temperature) but the virus was eventually eliminated. At 30 degrees C (the permissive temperature) in L cells all mutants established persistent infection. In ME cells at 30 degrees C, RNA- mutants (unable to synthesize virus-specified RNA at 39 degrees C) established persistent infection whereas the cells did not recover from infection with RNA+ mutants (able to synthesize virus-specified RNA at 39 degrees C). The wt virus was less cytopathic in L cells than in BHK or ME cells. Interferon was produced by both L and ME cells at 30 degrees C and 39 degrees C, but its activity could not be detected in either cell line at 30 degrees C. It is proposed that establishment of persistent infection is dependent on reduced cytopathogenicity in the early stage of infection, and that further evolution of the virus then occurs to a less cytopathic form. Elimination of the virus at 39 degrees C is probably due to the action of interferon.

AU - Sawicki SG, Sawicki DL, Kaariainen L, Keranen S
TI - A Sindbis virus mutant temperature-sensitive in the regulation of minus-strand RNA synthesis.
SO - Virology 1981 Nov;115(1):161-72

AU - Stollar V, Malinoski F
TI - The effects of adenosine and guanosine on the replication of Sindbis and vesicular stomatitis viruses in Aedes albopictus cells.
SO - Virology 1981 Nov;115(1):57-66

AU - Rupprecht KM, Sonenberg N, Shatkin AJ, Hecht SM
TI - Design and preparation of affinity columns for the purification of eukaryotic messenger ribonucleic acid cap binding protein.
SO - Biochemistry 1981 Nov 10;20(23):6570-7
AB - 2',3'-O-[1-(2-Carboxyethyl) ethylidene]-7-methylguanosine 5'-diphosphate (5) and 7-(5-carboxypentyl) guanosine 5'-diphosphate (13) have been synthesized and immobilized on AH-Sepharose 4B to the extent of 17.4 and 36.6 mumol of ligand/g of gel, respectively. The affinity resins thus derives were employed in columns for the purificaton of 24K cap binding protein (CBP) from rabbit reticulocytes. Each resin was found to retain the protein of interest; elution of 24K CBP could then be effected by washing with 70 microM m7GDP. The 24K CBPs released from both columns were found to be active, both as judged by a cross-linking assay that utilized 10(4)-oxidized methyl-3H-labeled reovirus mRNA as a substrate for the protein and also by the ability of the isolated 24K CBP to stimulate the translocation of capped Sindbis virus mRNA in HeLa cell extracts.

AU - Kowal KJ, Stollar V
TI - Temperature-sensitive host-dependent mutants of Sindbis virus.
SO - Virology 1981 Oct 15;114(1):140-8

AU - Collins PL, Fuller FJ, Marcus PI, Hightower LE, Ball LA
TI - Synthesis and processing of Sindbis virus nonstructural proteins in vitro.
SO - Virology 1982 Apr 30;118(2):363-79

AU - Erwin C, Brown DT
TI - Requirement of cell nucleus for Sindbis virus replication in cultured Aedes albopictus cells.
SO - J Virol 1983 Feb;45(2):792-9
AB - The ability of Sindbis virus to grow in enucleated BHK-21 (vertebrate) and Aedes albopictus (invertebrate) cells was tested to determine the dependence of this virus upon nuclear function in these two phylogenetically unrelated hosts. Although both cell types could be demonstrated to produce viable cytoplasts (enucleated cells) which produced virus-specific antigen subsequent to infection. BHK cytoplasts produced a significant number of progeny virions, whereas mosquito cytoplasts did not. The production of vesicular stomatitis virus in mosquito cells was not significantly reduced by enucleation. That such a host function was not essential for vesicular stomatitis virus growth in insect cells is supported by the observation that the production of this virus by mosquito cells is not actinomycin D sensitive. This result agrees with a previously published report in which it was shown that Sindbis virus maturation in invertebrate cells is inhibited by actinomycin D, indicating a possible requirement for host cell nuclear function (Scheefers-Borchel et al., Virology, 110:292-301, 1981).

AU - Baric RS, Carlin LJ, Johnston RE
TI - Requirement for host transcription in the replication of Sindbis virus.
SO - J Virol 1983 Jan;45(1):200-5
AB - Host cell involvement in Sindbis virus (SB) replication was examined in cells which had been treated with either actinomycin D (AMD) or alpha-amanitin (alpha-A). Treatment with these inhibitors of host transcription before infection reduced the ability of cells to support SB growth by 1 to 2 orders of magnitude, while having little or no effect on the replication of vesicular stomatitis virus. SB replication was sensitive to alpha-A in wild-type Chinese hamster ovary (CHO) cells but was resistant to alpha-A in CHOama-1 cells, a line which contains an alpha-A-resistant RNA polymerase II. A mutant of SB, SBamr, was isolated by mutagenesis followed by selection in cells which had been treated with AMD. SBamr grew normally not only in cells treated with AMD but also in alpha-A-treated cells. Our results suggest (i) that the synthesis of cellular mRNA (and presumably protein) is required for replication of SB, (ii) that prior treatment with either drug affects the same aspect of SB replication, and (iii) that mutations in the SB genome allow the virus to overcome the effect of inhibitors of host transcription.

AU - Baric RS, Lineberger DW, Johnston RE
TI - Reduced synthesis of Sindbis virus negative strand RNA in cultures treated with host transcription inhibitors.
SO - J Virol 1983 Jul;47(1):46-54
AB - Host cell involvement in Sindbis virus (SB) RNA synthesis was examined in cells which had been treated before infection with actinomycin D or alpha-amanitin (alpha-A). Overall synthesis of SB RNA was reduced significantly in CHO cells treated for 18 h before infection with alpha-A. However, SB RNA was produced at near normal levels in CHOama-1 cells, a line which contains an alpha-A-resistant RNA polymerase II. In BHK or CHO cells infected with SBamr, a mutant which replicates normally in cells pretreated with either actinomycin D or alpha-A, viral RNA synthesis was not decreased. The levels of negative strand RNA and of replicative forms I, II, and III in SB-infected cells were progressively reduced with increasing times of pretreatment with host transcription inhibitors, indicating fewer functional replicative intermediates in treated cells. Replicative events after replicative intermediate formation also were inhibited but only to the extent predicted by the reduction in replicative intermediates. Similarly, events preceding negative strand synthesis, adsorption, penetration, uncoating, and translation of nonstructural proteins, apparently were not impeded in treated cells. Therefore, our results are consistent with the involvement of a host component after translation of the nonstructural proteins but before or during the synthesis of SB negative strand RNA.

AU - Strauss EG, Tsukeda H, Simizu B
TI - Mutants of sindbis virus. IV. Heterotypic complementation and phenotypic mixing between temperature-sensitive mutants and wild-type Sindbis and Western equine encephalitis viruses.
SO - J Gen Virol 1983 Jul;64 (Pt 7):1581-90
AB - Heterotypic complementation between temperature-sensitive (ts) mutants of Sindbis (SIN) and Western equine encephalitis (WEE) viruses occurs under appropriate conditions. One heterotypic pair, SIN ts153 X WEE ts39 showed efficient complementation, and four other combinations gave detectable complementation, indicating that these two viruses, which are closely related serologically and biochemically, are sufficiently closely related to complement each other functionally. Cells mixedly infected with ts mutants or wild-type strains of both SIN and WEE viruses produced phenotypically mixed virions, in addition to both parental viruses. Various types of phenotypically mixed virions have been identified by neutralization with corresponding antisera, by thermal inactivation and by temperature sensitivity of replication. Some virions contained WEE genomes and envelopes containing primarily SIN proteins. Other phenotypically mixed virus yields contained primarily doubly neutralizable viruses which are presumed to have a mosaic of envelope proteins. Phenotypically mixed virions were morphologically indistinguishable from the parental types.

AU - Ghubril VA
TI - Blockage of antiviral induction of interferon by homologous cell biochemical activity: effect of chicken embryo fibroblast mitotic cell cycle phases on Sindbis virus growth.
SO - J Virol 1983 Sep;47(3):637-41
AB - The antiviral activity of interferon, measured as the reduction of viral yield, was studied as a function of the cell cycle phases. The present study shows that cells which are about to enter DNA replication phase S and cells that are in mitosis phase M are not refractive to viral infection when treated with interferon. The growth of Sindbis virus, used as the challenger, dropped considerably at the G1-S junction, at mitosis phase M, and as cells entered into a deeper quiescent state.

AU - Garry RF, Ulug ET, Bose HR Jr
TI - Induction of stress proteins in Sindbis virus- and vesicular stomatitis virus-infected cells.
SO - Virology 1983 Sep;129(2):319-32
AB - Two proteins which are related to certain proteins induced by hyperthermia (heat shock proteins; hsp) are synthesized during lytic infection of chick embryo (CE) cells by Sindbis virus or vesicular stomatitis virus (VSV). Incubation of the infected cells at elevated temperature further increased the rate of synthesis of these proteins. The stress proteins induced by Sindbis virus had different mobilities on SDS-polyacrylamide gels compared to related stress proteins induced in mock-infected CE cells. Induction of the stress proteins in Sindbis virus- and VSV-infected CE cells was actinomycin D sensitive. Kinetic studies indicated that induction of the stress proteins is an early event during infection. The lytic virus-induced selective termination of host protein synthesis did not affect the synthesis of these proteins. Furthermore, the synthesis of these virus-induced stress proteins was resistant, relative to the synthesis of most host proteins, to alterations in the intracellular concentrations of Na+ and K+. The synthesis of a protein related to a major low-molecular-weight hsp of CE cells was not induced after Sindbis virus or VSV infection. Immunoprecipitation experiments and sedimentation analyses demonstrated that significant levels of the capsid protein (C) of Sindbis virus and nucleocapsid protein (N) of VSV are physically associated with a hsp in lysates of infected CE cells.

AU - Strauss EG, Rice CM, Strauss JH
TI - Sequence coding for the alphavirus nonstructural proteins is interrupted by an opal termination codon.
SO - Proc Natl Acad Sci U S A 1983 Sep;80(17):5271-5
AB - We have obtained the nucleotide sequence of the genomic RNAs of two alphaviruses, Sindbis virus and Middelburg virus, over an extensive region encoding the nonstructural (replicase) proteins. In both viruses in an equivalent position an opal (UGA) termination codon punctuates a long otherwise open reading frame. The nonstructural proteins are translated as polyprotein precursors that are processed by posttranslational cleavage into four polypeptide chains; the sequence data presented here indicate that the COOH-terminal polypeptide, ns72, may be produced by read-through of this opal codon. The high degree of amino acid homology between the ns72 polypeptides of the two viruses, in contrast to the lack of conserved sequence upstream from the read-through site, suggests that ns72 plays an important role in viral replication, possibly modulating the action of other replicase components. MS - GENBANK/J02246, GENBANK/J02363, GENBANK/J02364, GENBANK/J02365, GENBANK/J02366, GENBANK/J02367, GENBANK/V00073

AU - Stollar V, Hardy JL
TI - Host-dependent mutants of Sindbis virus whose growth is restricted in cultured Aedes albopictus cells produce normal yields of virus in intact mosquitoes.
SO - Virology 1984 Apr 15;134(1):177-83
AB - Two host-dependent (hd), temperature-sensitive (ts) mutants of Sindbis virus (SV), clones 35 and 58, which are restricted in their ability to grow at 34 degrees in cultured Aedes albopictus cells [K.J. Kowal and V. Stollar (1981). Virology 114, 140-148] were not restricted in their ability to grow at 28-34 degrees after intrathoracic inoculation of Ae. albopictus, Ae. dorsalis, Ae. epactius, Anopheles freeborni, and Culex tarsalis mosquitoes. Compared to standard SV (SVSTD), the growth of SV clone 35 was somewhat restricted in parenterally infected C. pipiens females. Titers of SVSTD and the two mutants were modulated to relative low levels by some Ae. albopictus and Ae. dorsalis females after 192 hr incubation at 34 degrees but not at 28 degrees, suggesting that under some conditions mosquitoes have the ability to control the replication of SV. The hd and ts phenotypes of SV mutants were retained during growth at 31 degrees for 36 hr and 72 hr in Ae. epactius and Ae. dorsalis females, respectively, and at 28 degrees or 34 degrees for 192 hr in Ae. albopictus females. The demonstration that mutant viral clones, which are restricted in cultured mosquito cells, can produce normal yields of virus in intact mosquitoes indicates that considerable caution must be exercised in extrapolating results obtained with cultured cells to the level of the intact organism.

AU - Strauss EG, Rice CM, Strauss JH
TI - Complete nucleotide sequence of the genomic RNA of Sindbis virus.
SO - Virology 1984 Feb;133(1):92-110
AB - The entire nucleotide sequence of the genomic RNA of the type virus of the alphavirus genus, Sindbis virus, has been determined. The genome is 11,703 nucleotides in length, exclusive of the 5' cap and the 3'-terminal poly(A) tract. After the 5'-terminal cap there are 59 nucleotides of 5' nontranslated nucleic acid followed by a reading frame of 7539 nucleotides that encodes the nonstructural polypeptides and which is open except for a single opal termination codon. Following 48 untranslated bases located in the junction region which separates the nonstructural and structural protein coding sequences, there is an open reading frame 3735 nucleotides long that encodes the structural proteins. Finally, the 3' untranslated region is 322 nucleotides long. The nonstructural proteins are translated from the genomic RNA as two polyprotein precursors. The first is 1896 amino acids in length and terminates at an opal codon at position 1897. This polyprotein is processed to produce three polypeptides called nsP1, nsP2, and nsP3. Sites of post-translational cleavage to produce these three proteins have been tentatively located using available N-terminal amino acid sequence data. In both cases cleavage probably occurs between the two alanine residues in the sequence Gly-Ala-Ala. The fourth nonstructural protein, nsP4, is produced when readthrough of the opal codon produces a second polyprotein precursor of length 2513 amino acids, which is also cleaved posttranslationally. The structural proteins are translated from a subgenomic message which begins at nucleotide 7598, is 4106 nucleotides in length (exclusive of the poly(A) tract), and is coterminal with the 3' end of the genomic RNA. The structural proteins are also translated as a polyprotein precursor which is cleaved to produce a nucleocapsid protein and two integral membrane glycoproteins as well as two small peptides not present in the mature virion. A replication strategy for Sindbis virus based upon the complete nucleotide sequence, as well as prior data, is presented. MS - GENBANK/J02363, GENBANK/J02364, GENBANK/J02365, GENBANK/J02366, GENBANK/J02367, GENBANK/V00073

AU - Shablinskaia LM, Tsilinskii IaIa
TI - [Induction of alphavirus ts-mutations by N-methyl-N-nitro-N-nitrosoguanidine added to the replicating virus]. [RUSSIAN]
SO - Vopr Virusol 1984 Jan-Feb;29(1):86-9
AB - The possibility of producing ts mutants of Sindbis and eastern equine encephalomyelitis (EEE) virus by treatment of replicating virus with N-methyl-N-nitro-N-nitrosoguanidine was studied. N-methyl-N-nitro-N-nitrosoguanidine added in a concentration of 20 micrograms/ml to chick fibroblast cultures infected with Sindbis virus for 4 hours was shown to induce ts mutations in the virus. Under similar conditions no ts mutants of EEE virus could be obtained. The content of ts mutants in the mutagenized populations of Sindbis virus was 7.9%. Eight ts mutants were isolated. Their temperature-sensitive defect was expressed to various degrees. The plaque-forming efficiency at 40 degrees C/35 degrees C ranged from 6.3 X 10(-2) to 1.7 X 10(-8), and the yield from 5.6 X 10(-2) to 2.8 X 10(-4).

AU - Haseloff J, Goelet P, Zimmern D, Ahlquist P, Dasgupta R, Kaesberg P
TI - Striking similarities in amino acid sequence among nonstructural proteins encoded by RNA viruses that have dissimilar genomic organization.
SO - Proc Natl Acad Sci U S A 1984 Jul;81(14):4358-62
AB - The plant viruses alfalfa mosaic virus (AMV) and brome mosaic virus (BMV) each divide their genetic information among three RNAs while tobacco mosaic virus (TMV) contains a single genomic RNA. Amino acid sequence comparisons suggest that the single proteins encoded by AMV RNA 1 and BMV RNA 1 and by AMV RNA 2 and BMV RNA 2 are related to the NH2-terminal two-thirds and the COOH-terminal one-third, respectively, of the largest protein encoded by TMV. Separating these two domains in the TMV RNA sequence is an amber termination codon, whose partial suppression allows translation of the downstream domain. Many of the residues that the TMV read-through domain and the segmented plant viruses have in common are also conserved in a read-through domain found in the nonstructural polyprotein of the animal alphaviruses Sindbis and Middelburg. We suggest that, despite substantial differences in gene organization and expression, all of these viruses use related proteins for common functions in RNA replication. Reassortment of functional modules of coding and regulatory sequence from preexisting viral or cellular sources, perhaps via RNA recombination, may be an important mechanism in RNA virus evolution.

AU - Barrett AD, Dimmock NJ
TI - Variation in homotypic and heterotypic interference by defective interfering viruses derived from different strains of Semliki Forest virus and from Sindbis virus.
SO - J Gen Virol 1984 Jun;65 ( Pt 6):1119-22
AB - There was strong interference between various virulent and avirulent strains of Semliki Forest virus (SFV) and their respective defective interfering (DI) viruses but in other combinations interference was variable: it could be equally strong, weak or could not be demonstrated. On passage, this spectrum of interfering activity changed, some combinations showing greater interference than before and others less. Heterotypic interference between DI SFV, DI Sindbis virus and standard viruses was clearly demonstrated although this was strongest between DI SFV preparations and Sindbis standard virus than in the reciprocal combinations. Variation in interference between DI SFVs and different SFV strains was similar in magnitude to that between DI SFVs and Sindbis virus, suggesting that a similar DI RNA sequence is recognized by both viruses.

AU - Theilmann D, Eaton BT, Downe AE
TI - Properties of Sindbis virus variants from infected Culex tarsalis mosquitoes.
SO - J Gen Virol 1984 May;65 ( Pt 5):945-53
AB - Plaque-purified Sindbis virus was passed three times in Culex tarsalis mosquitoes and progeny viruses were isolated by plaque purification on a cloned line of Aedes albopictus cells. Nine of ten clones examined differed from wild-type (wt) virus with respect to their plaque morphology characteristics in chick embryo fibroblast (CEF) and/or A. albopictus cells. Seven clones were temperature-sensitive and failed to replicate or synthesize viral RNA in CEF cells at 41 degrees C. At 35 degrees C in CEF cells the majority of isolates synthesized less viral RNA than wt virus. In contrast, all cloned isolates synthesized viral RNA more rapidly than wild-type virus in A. albopictus cells.

AU - Kamer G, Argos P
TI - Primary structural comparison of RNA-dependent polymerases from plant, animal and bacterial viruses.
SO - Nucleic Acids Res 1984 Sep 25;12(18):7269-82
AB - Possible alignments for portions of the genomic codons in eight different plant and animal viruses are presented: tobacco mosaic, brome mosaic, alfalfa mosaic, sindbis, foot-and-mouth disease, polio, encephalomyocarditis, and cowpea mosaic viruses. Since in one of the viruses (polio) the aligned sequence has been identified as an RNA-dependent polymerase, this would imply the identification of the polymerases in the other viruses. A conserved fourteen-residue segment consisting of an Asp-Asp sequence flanked by hydrophobic residues has also been found in retroviral reverse transcriptases, a bacteriophage, influenza virus, cauliflower mosaic virus and hepatitis B virus, suggesting this span as a possible active site or nucleic acid recognition region for the polymerases. Evolutionary implications are discussed.

AU - Tsiang M, Monroe SS, Schlesinger S
TI - Studies of defective interfering RNAs of Sindbis virus with and without tRNAAsp sequences at their 5' termini.
SO - J Virol 1985 Apr;54(1):38-44
AB - Three of six independently derived defective interfering (DI) particles of Sindbis virus generated by high-multiplicity passaging in cultured cells have tRNAAsp sequences at the 5' terminus of their RNAs (Monroe and Schlesinger, J. Virol. 49:865-872, 1984). In the present work, we found that the 5'-terminal sequences of the three tRNAAsp-negative DI RNAs were all derived from viral genomic RNA. One DI RNA sample had the same 5'-terminal sequence as the standard genome. The DI RNAs from another DI particle preparation were heterogeneous at the 5' terminus, with the sequence being either that of the standard 5' end or rearrangements of regions near the 5' end. The sequence of the 5' terminus of the third DI RNA sample consisted of the 5' terminus of the subgenomic 26S mRNA with a deletion from nucleotides 24 to 67 of the 26S RNA sequence. These data showed that the 5'-terminal nucleotides can undergo extensive variations and that the RNA is still replicated by virus-specific enzymes. DI RNAs of Sindbis virus evolve from larger to smaller species. In the two cases in which we followed the evolution of DI RNAs, the appearance of tRNAAsp-positive molecules occurred at the same time as did the emergence of the smaller species of DI RNAs. In pairwise competition experiments, one of the tRNAAsp-positive DI RNAs proved to be the most effective DI RNA, but under identical conditions, a second tRNAAsp-positive DI RNA was unable to compete with the tRNAAsp-negative DIs. Therefore, the tRNAAsp sequence at the 5' terminus of a Sindbis DI RNA is not the primary factor in determining which DI RNA becomes the predominant species in a population of DI RNA molecules. MS - GENBANK/K02741

AU - Ahlquist P, Strauss EG, Rice CM, Strauss JH, Haseloff J, Zimmern D
TI - Sindbis virus proteins nsP1 and nsP2 contain homology to nonstructural proteins from several RNA plant viruses.
SO - J Virol 1985 Feb;53(2):536-42
AB - Although the genetic organization of tobacco mosaic virus (TMV) differs considerably from that of the tripartite viruses (alfalfa mosaic virus [AlMV] and brome mosaic virus [BMV]), all of these RNA plant viruses share three domains of homology among their nonstructural proteins. One such domain, common to the AlMV and BMV 2a proteins and the readthrough portion of TMV p183, is also homologous to the readthrough protein nsP4 of Sindbis virus (Haseloff et al., Proc. Natl. Acad. Sci. U.S.A. 81:4358-4362, 1984). Two more domains are conserved among the AlMV and BMV 1a proteins and TMV p126. We show here that these domains have homology with portions of the Sindbis proteins nsP1 and nsP2, respectively. These results strengthen the view that the four viruses share mechanistic similarities in their replication strategies and may be evolutionarily related. These results also suggest that either the AlMV 1a, BMV 1a, and TMV p126 proteins are multifunctional or Sindbis proteins nsP1 and nsP2 function together as subunits in a single complex.

AU - Sawicki DL, Sawicki SG
TI - Functional analysis of the A complementation group mutants of Sindbis HR virus.
SO - Virology 1985 Jul 15;144(1):20-34
AB - The 10 members of the A complementation group of temperature-sensitive (ts) mutants of SIN HR, the heat-resistant strain of Sindbis virus, were divided into two phenotypic subgroups. Subgroup I mutants (ts15, ts17, ts21, ts24, and ts133) demonstrated temperature-sensitive 26 S mRNA synthesis, whereas subgroup II mutants (ts4, ts14, ts16, ts19, and ts138) did not; both ts4 and ts19 demonstrated defective 26 S mRNA synthesis at 30 degrees. None of the A group mutants demonstrated temperature-sensitive 49 S plus-strand synthesis. Temperature-sensitive cleavage of ns230 was demonstrated by subgroup I mutants, except ts21, but not by subgroup II mutants. A revertant of ts133 that grew at 40 degrees retained temperature-sensitive 26 S mRNA synthesis but lost temperature-sensitive cleavage of ns230 and the RNA-negative phenotype. Only ts4, like ts11 of the B complementation group, demonstrated temperature-sensitive minus-strand RNA synthesis. In addition to ts24, cells infected with ts17 or ts133 continued to synthesize minus strands after shiftup in the absence of continued protein synthesis, and resumed synthesis of minus strands if shifted to the nonpermissive temperature after minus-strand synthesis had ceased at the permissive temperature.

AU - Durbin RK, Stollar V
TI - Sindbis virus mutants able to replicate in methionine-deprived Aedes albopictus cells.
SO - Virology 1985 Jul 30;144(2):529-33
AB - Previous work from this laboratory has shown that the replication of Sindbis virus (SV) in Aedes albopictus cells is sensitive to methionine withdrawal. This sensitivity is thought to reflect a diminished concentration of S-adenosylmethionine (Ado Met) resulting from methionine starvation. Serial passage of SV on Ae. albopictus cells maintained in low concentrations of methionine gave rise to a population of mutants whose replication in mosquito cells was resistant to methionine starvation. In vertebrate cells, these mutants were also resistant to inhibition by cycloleucine. We favor the hypothesis that the adaptation to low methionine reflects the accumulation of mutations resulting in a viral RNA "cap" methyltransferase with an increased affinity for Ado Met.

AU - Lopez S, Bell JR, Strauss EG, Strauss JH
TI - The nonstructural proteins of Sindbis virus as studied with an antibody specific for the C terminus of the nonstructural readthrough polyprotein.
SO - Virology 1985 Mar;141(2):235-47
AB - A dodecapeptide containing the sequence of the C terminus of the nonstructural polyprotein of Sindbis virus has been synthesized and used to immunize rabbits. The antisera obtained precipitated polypeptides from cells infected with the HR strains of Sindbis or with temperature-sensitive mutants ts11 or ts18. Four different polypeptides, having apparent molecular weights of approximately 250,000, 220,000, 155,000, and 72,000, were immunoprecipitated by the antipeptide antiserum. The largest of these polypeptides is sufficiently large to represent a polyprotein translated from the entire nonstructural region of the genome. These data suggest that nsP4 of molecular weight 72,000 is produced by translation of the entire nonstructural region of the genome, which requires readthrough of an opal termination codon immediately upstream of nsP4, followed by post-translational cleavage of this polyprotein. The amounts of nsP4 and its precursors found in infected cells are small relative to the amounts of other nonstructural proteins present, as would be expected if readthrough of a termination codon is required. In addition, the relative amounts of nsP4 and of its precursors differ in HR-infected or ts mutant-infected cells and differ with temperature of infection, suggesting that temperature of infection or ts lesions affect translation and processing of the precursor polyprotein.

AU - Adams RH, Brown DT
TI - BHK cells expressing Sindbis virus-induced homologous interference allow the translation of nonstructural genes of superinfecting virus.
SO - J Virol 1985 May;54(2):351-7
AB - The process by which Sindbis virus excludes superinfecting homologous virus was investigated with the use of temperature-sensitive mutants. Mutants in two RNA-negative complementation groups were found to be defective in their ability to establish interference at the nonpermissive temperature. These mutants were unable to establish interference in a mixed infection (complementation), suggesting that both were defective in a common gene product. Homologous interference was found to block the replication of superinfecting virus after attachment, penetration, and translation of the nonstructural genes encoded in the virus RNA. The production of nonstructural gene products of superinfecting wild-type virus was found to enhance the replication of certain RNA- temperature-sensitive interfering viruses at the permissive and the nonpermissive temperature. The ability of certain RNA- mutants to establish homologous interference and to demonstrate enhanced growth after superinfection with wild-type virus was interpreted to produce a model implicating both virus and host components in the establishment of homologous interference and in the replication of Sindbis virus RNA.

AU - Condreay LD, Brown DT
TI - Exclusion of superinfecting homologous virus by Sindbis virus-infected Aedes albopictus (mosquito) cells.
SO - J Virol 1986 Apr;58(1):81-6
AB - The infection of tissue-cultured Aedes albopictus (mosquito) cells by an alphavirus ultimately results in a persistently infected cell population which can be maintained in the laboratory for years. One characteristic of this culture is that it will not support the replication of superinfecting homologous virus. We have previously shown that mosquito cells persistently infected with Sindbis virus produce an antiviral agent which when applied to uninfected mosquito cells suppresses Sindbis virus replication. The exclusion of virus replication in the antiviral-agent-treated cells is similar to the phenomenon of homologous interference described in alphavirus-infected vertebrate cells. In this study we examined the expression of homologous interference in three lines of mosquito cells and compared the expression of homologous interference to the effects of the antiviral activity. The cell lines were found to differ in their ability to express homologous interference, and evidence suggests that the mosquito cells may suppress replication by homologous interference or by the action of the antiviral agent.

AU - Tatem J, Stollar V
TI - Dominance of the CPE(+) phenotype in hybrid Aedes albopictus cells infected with Sindbis virus.
SO - Virus Res 1986 Aug;5(2-3):121-30
AB - The effect of Sindbis virus (SV) infection was analyzed in hybrid Aedes albopictus cells formed by fusing ouabain-resistant CPE(+) cells to CPE(-) alpha-amanitin resistant cells. Although the 24-h yields of virus from the parental CPE(+) and CPE(-) clones were similar, the rates of viral RNA synthesis and virus release at early times post-infection were higher in the CPE(+) cells. In all eight hybrid clones studied, the CPE(+) phenotype was dominant. In addition, the kinetics of viral RNA synthesis and virus release in the hybrids closely resembled what was observed in the CPE(+) parent clone. These data indicate that both in the parental and in the hybrid cells, expression of SV-induced CPE is associated with a high level of viral RNA synthesis and a rapid production of virus during the early period after infection. It is suggested that CPE(+) cells contain some factor or activity which is lacking or less abundant in the CPE(-) cells and that this activity is important in the regulation of viral RNA synthesis.

AU - Garry RF, Bostick DA, Ulug ET
TI - Sindbis virus infection increases hexose transport in quiescent cells.
SO - Virology 1986 Dec;155(2):378-91
AB - Sindbis virus infection of baby hamster kidney cells or chick embryo cells resulted in a significant increase in the rate of uptake of [2-3H]deoxy-D-glucose ([3H]dGlu). Stimulation of hexose transport in Sindbis virus-infected cells occurred only if the cells were rendered quiescent by culturing at high density or by serum starvation. In contrast, Sindbis virus-induced inhibition of potassium transport, measured as a decrease in the uptake of 86Rb+, was independent of cell growth state. Stimulation of [3H]dGlu uptake in Sindbis virus-infected cells was the result of an increase in the Vmax of the hexose transporter, but not a change in the Km. The stimulation of [3H]dGlu uptake induced by Sindbis virus was insensitive to the drug actinomycin D, but was blocked by cordycepin. The stimulation was also insensitive to treatment with tunicamycin, which prevented the virally induced inhibition of the plasma membrane-associated Na+/K+ ATPase and termination of host protein synthesis.

AU - Kuge S, Saito I, Nomoto A
TI - Primary structure of poliovirus defective-interfering particle genomes and possible generation mechanisms of the particles.
SO - J Mol Biol 1986 Dec 5;192(3):473-87
AB - The genomes of defective-interfering (DI) particles derived from the Sabin strain of type 1 poliovirus (PV1(Sab] were characterized by nuclease S1 mapping using complementary DNA (cDNA) copies of PV1(Sab) genome as probes. The results demonstrated variety in the size and location of the deletions, which were compatible with our previous prediction. The results further indicated that the locations of the deletions were limited within the internal genome region encoding viral capsid proteins and that the deletion sites were clustered in certain areas on the genome. Sequence analysis of a number of cloned cDNAs to the DI genomes revealed that every DI genome retained the correct reading frame for viral protein synthesis. These results strongly suggested that one or all of the viral non-structural proteins might be cis-acting at least at a certain stage in viral replication. A computer search for secondary structures with regard to the deletion sites provided a possible common structure from which, supported by sequences existing on the plus or minus RNA strand of PV1(Sab), deletion regions looped out from the remaining sequences. Replicase might, therefore, skip these transiently formed loop structures with certain frequencies, resulting in the generation of DI genomes. This model could also be considered as a model for genetic recombination in these RNA genomes. Possible "supporting sequences" were also found for every rearranged site on the RNAs of influenza virus and sindbis virus. Thus, we propose a new copy-choice model, designated the "supporting sequence-loop model", for the generation of rearrangements occurring on single-stranded RNA genomes.

AU - Levis R, Weiss BG, Tsiang M, Huang H, Schlesinger S
TI - Deletion mapping of Sindbis virus DI RNAs derived from cDNAs defines the sequences essential for replication and packaging.
SO - Cell 1986 Jan 17;44(1):137-45
AB - Defective-interfering (DI) genomes of a virus contain sequence information essential for their replication and packaging. They need not contain any coding information and therefore are a valuable tool for identifying cis-acting, regulatory sequences in a viral genome. To identify these sequences in a DI genome of Sindbis virus, we cloned a cDNA copy of a complete DI genome directly downstream of the promoter for the SP6 bacteriophage DNA dependent RNA polymerase. The cDNA was transcribed into RNA, which was transfected into chicken embryo fibroblasts in the presence of helper Sindbis virus. After one to two passages the DI RNA became the major viral RNA species in infected cells. Data from a series of deletions covering the entire DI genome show that only sequences in the 162 nucleotide region at the 5' terminus and in the 19 nucleotide region at the 3' terminus are specifically required for replication and packaging of these genomes. MS - GENBANK/M12563

AU - Takkinen K
TI - Complete nucleotide sequence of the nonstructural protein genes of Semliki Forest virus.
SO - Nucleic Acids Res 1986 Jul 25;14(14):5667-82
AB - The nucleotide sequence coding for the nonstructural proteins of Semliki Forest virus has been determined from cDNA clones. The total length of this region is 7381 nucleotides, it contains an open reading frame starting at position 86 and ending at an UAA stop codon at position 7379-7381. This open reading frame codes for a 2431 amino acids long polyprotein, from which the individual nonstructural proteins are formed by proteolytic processing steps, so that nsPl is 537, nsP2 798, nsP3 482 and nsP4 614 amino acids. In the closely related Sindbis and Middelburg viruses there is an opal stop codon (UGA) between the genes for nsP3 and nsP4. Interestingly, no stop codon is found in frame in this region of the Semliki Forest virus 42S RNA. In other aspects the amino acid sequence homology between Sindbis, Middelburg and Semliki Forest virus nonstructural proteins is highly significant. MS - GENBANK/J02361, GENBANK/J02362, GENBANK/L00018, GENBANK/V01399, GENBANK/V01400, GENBANK/V01401, GENBANK/X04129

AU - Sawicki SG, Sawicki DL
TI - The effect of loss of regulation of minus-strand RNA synthesis on Sindbis virus replication.
SO - Virology 1986 Jun;151(2):339-49
AB - During the replication cycle of Sindbis virus minus-strand synthesis stops normally at the time that plus-strand synthesis reaches a maximum rate. We have isolated and characterized revertants of ts24, a member of the A complementation group of Sindbis HR mutants, that we had demonstrated previously to have a temperature-sensitive defect in the regulation of minus-strand synthesis. These revertants of ts24 replicated efficiently at 40 degrees but nevertheless retained the temperature sensitive defect in the regulation of minus-strand synthesis: they continued to synthesize minus strands late in the replication cycle at 40 degrees but not at 30 degrees and in the presence or absence of protein synthesis. Although failure to regulate the synthesis of minus strands resulted in continuous minus-strand synthesis and in the accumulation of minus strands, the rate of plus-strand synthesis was not increased concertedly. Minus strands synthesized after the rate of plus-strand synthesis had become constant were demonstrated to be utilized as templates for 26 S mRNA synthesis. Thus, the change from an increasing to a constant rate of plus-strand synthesis during the alphavirus replication cycle cannot be governed solely by the number of minus strands that accumulate in infected cells. We present a model for the preferential utilization of minus strands as a mechanism for the cessation of minus-strand synthesis that occurs normally during alphavirus replication.

AU - McClure MA, Perrault J
TI - RNA virus genomes hybridize to cellular rRNAs and to each other.
SO - J Virol 1986 Mar;57(3):917-21
AB - In this communication we show that the RNA genomes of vesicular stomatitis, Sindbis, and reoviruses can specifically hybridize under stringent conditions to the large rRNAs present in HeLa cell cytoplasmic extracts. In addition, we show that some virus genome RNAs can also hybridize to each other. On the basis of our previous detailed studies identifying specific regions of hybridization between the poliovirus genome and 28S rRNA, we suggest that a similar phenomenon of "patchy complementary" may be responsible for the interactions described here (M. A. McClure and J. Perrault, Nucleic Acids Res. 13:6797-6816, 1985). The possible biological implications of these cross-reacting hybridizations and practical considerations in the use of viral probes for diagnosis are discussed.

AU - Kaariainen L, Takkinen K, Keranen S, Soderlund H
TI - Replication of the genome of alphaviruses.
SO - J Cell Sci Suppl 1987;7:231-50
AB - The genome of Semliki Forest virus (SFV) is 11,442 nucleotides with a 5' cap-structure and a 3' poly(A) tail of about 100 residues. The genome of the closely relate Sindbis virus (SIN) is slightly longer (11,703 nucleotides). The parental RNA is first translated from the 5' two thirds to yield; nsP1, nsP2, nsP3 and nsP4, which are cleaved from a polyprotein of 2431 amino acids (SFV). The parental genome is copied to a full-length minus strand with poly(U) at the 5' end. The minus strand is used as template for the synthesis of 42 S RNA in membrane-bound replicative-intermediate (RI) structures. In addition to 42 S RNA, a 3'-coterminal subgenomic 26 S mRNA, coding for the structural proteins, is synthesized by internal initiation at the minus strand. Capping and methylation of both plus-strand RNAs occur concomitantly with their synthesis. Analysis of Sindbis virus temperature-sensitive RNA-negative mutants have shown that one complementation group (B) is specifically associated with the synthesis of minus strands. Another, group F, is involved in the polymerization step of both minus- and plus-strand 42 S RNA, and of the 26 S mRNA. The synthesis of minus strands is normally dependent on protein synthesis. There is a shut off of the minus-strand RNA synthesis at about 3 h post-infection. This is apparently regulated by a virus-specific protein, represented by the complementation group A. The same protein is involved in the regulation of the initiation of 26 S RNA together with a component represented by group G mutants. Comparative analysis of SFV and SIN RNAs and DI RNAs of both viruses suggests that perhaps only 19 nucleotides from the 3' end and about 150 nucleotides from the 5' end are needed for replication of the alphavirus RNAs. In some SIN DI RNAs the proposed secondary structure at the 5' end is replaced by a cellular tRNA(ASP) suggesting that the secondary structure rather than nucleotide sequence is sufficient for the recognition by the viral polymerase. Even when the primary structure of the four non-structural proteins of both SFV and SIN is known, the correlation of the genetic data with the individual proteins has not yet been possible.

AU - Rice CM, Levis R, Strauss JH, Huang HV
TI - Production of infectious RNA transcripts from Sindbis virus cDNA clones: mapping of lethal mutations, rescue of a temperature-sensitive marker, and in vitro mutagenesis to generate defined mutants.
SO - J Virol 1987 Dec;61(12):3809-19
AB - We constructed full-length cDNA clones of Sindbis virus that can be transcribed in vitro by SP6 RNA polymerase to produce infectious genome-length transcripts. Viruses produced from in vitro transcripts are identical to Sindbis virus and show strain-specific phenotypes reflecting the source of RNA used for cDNA synthesis. The cDNA clones were used to confirm the mapping of the causal mutation of ts2 to the capsid protein. A general strategy for mapping Sindbis virus mutations is described and was used to identify two lethal mutations in an original full-length construct which did not produce infectious transcripts. An XbaI linker was inserted in the cDNA clone near the transcriptional start of the subgenomic mRNA; the resulting virus retains the XbaI recognition sequence, thus providing formal evidence that viruses are derived from in vitro transcripts of cDNA clones. The potential applications of the cDNA clones are discussed.

AU - Scheidel LM, Durbin RK, Stollar V
TI - Sindbis virus mutants resistant to mycophenolic acid and ribavirin.
SO - Virology 1987 May;158(1):1-7
AB - Previous work from this laboratory has demonstrated a correlation between the inhibition by ribavirin (Rbv), mycophenolic acid (MPA), or 2-amino thiadiazole (TDA) of Sindbis virus replication in Aedes albopictus mosquito cells and a reduction in cellular GTP levels. This reduction in GTP results from the inhibition by these drugs of inosine monophosphate dehydrogenase (IMPDH), the first enzyme specific for the de novo synthesis of GMP. By serial passage of SV in A. albopictus cells in the presence of 25 microM MPA, we have now isolated viral mutants which are highly resistant not only to MPA but also to Rbv and TDA. For example, whereas 500 microM Rbv reduced the plaquing efficiency of SVSTD by at least 10(6)-fold, the same concentration of Rbv reduced the plaquing efficiency of the MPA-resistant mutants less than 5-fold. This is the first example of a viral mutant resistant to the antiviral compound Rbv.

AU - Faragher SG, Meek AD, Rice CM, Dalgarno L
TI - Genome sequences of a mouse-avirulent and a mouse-virulent strain of Ross River virus.
SO - Virology 1988 Apr;163(2):509-26
AB - The nucleotide sequence of the genomic RNA of a mouse-avirulent strain of Ross River virus, RRV NB5092 (isolated in 1969), has been determined and the corresponding sequence for the prototype mouse-virulent strain, RRV T48 (isolated in 1959), has been completed. The RRV NB5092 genome is approximately 11,674 nucleotides in length, compared with 11,853 nucleotides for RRV T48. RRV NB5092 and RRV T48 have the same genome organization. For both viruses an untranslated region of 80 nucleotides at the 5' end of the genome is followed by a 7440-nucleotide open reading frame which is interrupted after 5586 nucleotides by a single opal termination codon. By homology with other alphaviruses, the 5586-nucleotide open reading frame encodes the nonstructural proteins nsP1, nsP2, and nsP3; a fourth nonstructural protein, nsP4, is produced by read-through of the opal codon. The RRV nonstructural proteins show strong homology with the corresponding proteins of Sindbis virus and Semliki Forest virus in terms of size, net charge, and hydropathy characteristics. However, homology is not uniform between or within the proteins; nsP1, nsP2, and nsP4 contain extended domains which are highly conserved between alphaviruses, while the C-terminal region of nsP3 shows little conservation in sequence or length between alphaviruses. An untranslated "junction" region of 44 nucleotides (for RRV NB5092) or 47 nucleotides (for RRV T48) separates the nonstructural and structural protein coding regions. The structural proteins (capsid-E3-E2-6K-E1) are translated from an open reading frame of 3762 nucleotides which is followed by a 3'-untranslated region of approximately 348 nucleotides (for RRV NB5092) or 524 nucleotides (for RRV T48). Excluding deletions and insertions, the genomes of RRV NB5092 and RRV T48 differ at 284 nucleotides, representing a sequence divergence of 2.38%. Sequence deletions or insertions were found only in the noncoding regions and include a 173-nucleotide deletion in the 3'-untranslated region of RRV NB5092, compared with RRV T48. In the coding regions, most of the nucleotide differences are silent; there are 36 amino acid differences in the nonstructural proteins and 12 in the structural proteins. The distribution of amino acid differences between the two RRV strains correlates with the location of domains which are poorly conserved in sequence between alphaviruses. The possible role of amino acid differences in envelope glycoproteins E1 and E2 in determining the different antigenic and biological properties of RRV NB5092 and RRV T48 is discussed. MS - GENBANK/M20162

AU - Condreay LD, Adams RH, Edwards J, Brown DT
TI - Effect of actinomycin D and cycloheximide on replication of Sindbis virus in Aedes albopictus (mosquito) cells.
SO - J Virol 1988 Aug;62(8):2629-35
AB - Production of Sindbis virus in the presence of transcription and translation inhibitors was examined in three Aedes albopictus cell lines. Addition of cycloheximide to heat-resistant Sindbis virus (SVHR)-infected mosquito cells arrested viral RNA synthesis completely, in contrast to the effects of this drug on virus-infected vertebrate cells. Production of mature virus by both SVHR (a variant commonly used as a wild-type virus) and SBamr (a mutant which is resistant to the effects of 18 h of pretreatment of vertebrate cells with actinomycin D) in mosquito u4.4, C6-36, and C7-10 cells was inhibited by 2 h of pretreatment with actinomycin D. Pretreatment with this drug for 2 h slightly enhances virus production in vertebrate cells. Treatment of mosquito cells with actinomycin D resulted in shutoff of SVHR RNA synthesis. The mutant SBamr was able to overcome the effects of actinomycin D on viral RNA synthesis and produced both 26S and 49S RNAs, even though no viral structural proteins or mature particles were produced in the presence of the drug. This result suggests that, in the presence of actinomycin, the nonstructural genes of SBamr are translated sufficiently to allow for RNA synthesis but that 26S RNA may not be translated to an extent that allows significant virus production. These data demonstrate that host components are involved in at least two distinct steps in the production of Sindbis virus in mosquito cells: (i) production of viral RNA and (ii) synthesis of viral structural polypeptides.

AU - Froshauer S, Kartenbeck J, Helenius A
TI - Alphavirus RNA replicase is located on the cytoplasmic surface of endosomes and lysosomes.
SO - J Cell Biol 1988 Dec;107(6 Pt 1):2075-86
AB - Using morphological and cell biological techniques, we have shown that the RNA replicase of Semliki Forest and Sindbis virus (two closely related alphaviruses) is located in complex ribonucleoprotein structures associated with the cytoplasmic surface of modified secondary lysosomes and endosomes. These nucleoprotein complexes often form a bridge between the membrane of the endocytic vacuole and the rough endoplasmic reticulum where the synthesis of the structural proteins of these viruses occurs. The results suggest that these cytopathic vacuoles constitute sites not only for viral RNA synthesis, but also for translation of structural proteins, and for the assembly of nucleocapsids.

AU - Tsiang M, Weiss BG, Schlesinger S
TI - Effects of 5'-terminal modifications on the biological activity of defective interfering RNAs of Sindbis virus.
SO - J Virol 1988 Jan;62(1):47-53
AB - We have been studying defective interfering (DI) genomes of the RNA enveloped virus Sindbis virus. Deletion mapping of a DI cDNA demonstrated that only sequences at the 3' and 5' termini of the genome are required for the DI RNA to be biologically active. We constructed a series of cDNAs that transcribe DI RNAs differing only in 5'-terminal sequences. Two of the 5' termini identical to ones found in naturally occurring DI RNAs are the 5' terminus of the virion RNA (DI-549) and the first 142 nucleotides from the 5' terminus of the subgenomic 26S mRNA attached to the 5' terminus of the virion RNA (DI-15). The latter has a 42-nucleotide deletion from nucleotides 25 to 66 in the 26S RNA sequence. These DI RNA transcripts were biologically active, but one (DI-526) which did not have the 42-nucleotide deletion of DI-15 was not replicated. The DI RNA isolated after the presumed amplification of the DI-526 transcript had deleted the first 54 nucleotides of the 26S RNA sequences. The 5' terminus of Sindbis virion RNA contains a stem and loop region that is conserved among alphaviruses. An 11-nucleotide deletion in DI-549 that disrupted this stem and loop rendered this DI RNA inactive. In contrast, this same deletion in DI-15 and one that removed an additional 100 nucleotides of the virion 5' terminus did not prevent its amplification. We did not detect by computer analysis any common secondary structures among the biologically active DI RNAs that distinguished them from those RNAs that were not amplified. Our results support the conclusion that tertiary structure or the ability of the RNA to adapt its structure upon interaction with protein is important in the recognition process.

AU - Condreay LD, Brown DT
TI - Suppression of RNA synthesis by a specific antiviral activity in Sindbis virus-infected Aedes albopictus cells.
SO - J Virol 1988 Jan;62(1):346-8
AB - An antiviral protein is released by mosquito cells persistently infected with Sindbis virus. Differences in both sensitivity to and production of this virus-specific activity were apparent in three independently produced Aedes albopictus cell lines. This activity inhibits total viral RNA synthesis in a time-dependent manner. The antiviral effect is maximally realized when cells are treated with the activity 48 h before infections. These data suggest that the antiviral activity induces an antiviral state in treated cells which prevents the formation or efficient function of viral RNA-synthesizing complexes.

AU - Morch MD, Boyer JC, Haenni AL
TI - Overlapping open reading frames revealed by complete nucleotide sequencing of turnip yellow mosaic virus genomic RNA.
SO - Nucleic Acids Res 1988 Jul 11;16(13):6157-73
AB - The complete nucleotide sequence of turnip yellow mosaic virus (TYMV) genomic RNA has been determined on a set of overlapping cDNA clones using a sequential sequencing strategy. The RNA is 6318 nucleotides long, excluding the cap structure. The genome organization deduced from the sequence confirms previous results of in vitro translation. A novel open reading frame (ORF) putatively encoding a Pro-rich and very basic 69K (K = kilodalton) protein is detected at the 5' end of the genome. It is initiated at the first AUG codon on the RNA and overlaps the major ORF that encodes the non structural 206K (previously referred to as 195K) protein of TYMV; its function is unknown. Several amino acid consensus sequences already described among plant and animal viruses are also found in the TYMV-encoded polypeptides. A comparison with other viruses whose RNA sequence is known leads to the conclusion that TYMV belongs to the "Sindbis-like" supergroup of viruses and could be related to Semliki forest virus. MS - GENBANK/X07441

AU - Durbin RK, De Clercq E, Stollar V
TI - SVLM21, a mutant of Sindbis virus able to grow in Aedes albopictus cells in the absence of methionine, shows increased sensitivity to S-adenosylhomocysteine hydrolase inhibitors such as neplanocin A.
SO - Virology 1988 Mar;163(1):218-21
AB - Inhibition of S-adenosylhomocysteine (AdoHcy) hydrolase by compounds such as Neplanocin A (NPA) leads to the build-up of AdoHcy and the inhibition of methyltransferase enzymes. Whether assayed by efficiency of plaquing or virus yield, SVLM21, a mutant of Sindbis virus resistant to methionine deprivation, was more sensitive to NPA than was the standard virus (SVstd) from which it was derived. For example, whereas 10 micrograms NPA/ml depressed the yield of SVLM21 by more than 30-fold, the yield of SVstd was not significantly affected. Similar differences in sensitivities were shown to three other compounds which inhibit AdoHcy hydrolase. These results support the idea that SVLM21 codes for an altered RNA methyltransferase.

AU - Hardy WR, Strauss JH
TI - Processing the nonstructural polyproteins of Sindbis virus: study of the kinetics in vivo by using monospecific antibodies.
SO - J Virol 1988 Mar;62(3):998-1007
AB - Plasmids were constructed which contained a large portion of each of the four nonstructural genes of Sindbis virus fused to the N-terminal two-thirds of the trpE gene of Escherichia coli. The large quantity of fusion protein induced from cells containing these plasmids was subsequently used as an antigen to generate polyclonal antisera in rabbits. Each antiserum was specific for the corresponding nonstructural protein and allowed ready identification of each nonstructural protein and of precursors containing the sequences of two or more nonstructural proteins. These antisera were used to determine the stability of the mature nonstructural proteins and to examine the kinetics of processing of the nonstructural proteins from their respective precursors in vivo. Pulse-chase experiments showed that the precursor P123 is cleaved with a half-life of approximately 19 min to produce P12 and nsP3; P12 is then cleaved with a half-life of approximately 9 min to produce nsP1 and nsP2. Thus, although the rate of cleavage between nsP1 and nsP2 is faster than that between nsP2 and nsP3, the latter cleavage must occur first and is therefore the rate-limiting step. The rate at which P34 is chased suggests that the cleavage between nsP3 and nsP4 is the last to occur; however the regulation of nsP4 function in Sindbis virus-infected cells may be even more complex than was previously thought. The products nsP1 and nsP2 (and nsP4) are relatively stable; nsP3, however, is unstable, with a half-life of about 1 h, and appears to be modified to produce heterodisperse, higher-molecular-mass forms. In general, the processing schemes used by Sindbis virus and Semliki Forest virus appear very similar, the major difference being that most nsP3 in Sindbis virus results from termination at an opal condon, whereas in Semliki Forest virus cleavage of the P34 precursor is required.

AU - Strauss EG, Levinson R, Rice CM, Dalrymple J, Strauss JH
TI - Nonstructural proteins nsP3 and nsP4 of Ross River and O'Nyong-nyong viruses: sequence and comparison with those of other alphaviruses.
SO - Virology 1988 May;164(1):265-74
AB - We have sequenced the nsP3 and nsP4 region of two alphaviruses, Ross River virus and O'Nyong-nyong virus, in order to examine these viruses for the presence or absence of an opal termination codon present between nsP3 and nsP4 in many alphaviruses. We found that Ross River virus possesses an in-phase opal termination codon between nsP3 and nsP4, whereas in O'Nyong-nyong virus this termination codon is replaced by an arginine codon. Previous studies have shown that two other alphaviruses, Sindbis virus and Middelburg virus, possess an opal termination codon separating nsP3 and nsP4 [E.G. Strauss, C.M. Rice, and J.H. Strauss (1983), Proc. Natl. Acad. Sci. USA 80, 5271-5275], whereas Semliki Forest virus possesses an arginine codon in lieu of the opal codon [K. Takkinen (1986), Nucleic Acids Res. 14, 5667-5682]. Thus, of the five alphaviruses examined to date, three possess the opal codon and two do not. Production of nsP4 requires readthrough of the opal codon in those alphaviruses that possess this termination codon and the function of the termination codon may be to regulate the amount of nsP4 produced. It is an open question then as to whether alphaviruses with no termination codon use other mechanisms to regulate the activity of this gene. The nsP4s of these five alphaviruses are highly conserved, sharing 71-76% amino acid sequence similarity, and all five contain the Gly-Asp-Asp motif found in many RNA virus replicases. The nsP3s are somewhat less conserved, sharing 52-73% amino acid sequence similarity throughout most of the protein, but each possesses a nonconserved C-terminal domain of 134 to 246 amino acids of unknown function. MS - GENBANK/M20303, GENBANK/M20539

AU - Skryabin KG, Morozov SYu, Kraev AS, Rozanov MN, Chernov BK, Lukasheva LI, Atabekov JG
TI - Conserved and variable elements in RNA genomes of potexviruses.
SO - FEBS Lett 1988 Nov 21;240(1-2):33-40
AB - The nucleotide sequences of genomic RNAs and predicted amino acid sequences of two strains of potato virus X and white clover mosaic potexvirus were compared to each other, and the proteins of different plus-RNA-containing plant viruses. The predicted non-virion proteins of potexviruses have direct sequence homology and common structural peculiarities with those of several 'Sindbis-like' plant viruses. The most conserved amino acid sequences were found to be located in the polypeptide encoded by the long 5'-proximal open reading frame (ORF1). The putative polypeptide encoded by the ORF2 starting beyond the ORF1 stop codon is clearly related to the presumptive NTP-binding domain of the ORF1-coded polypeptide. These results suggest possible functions for all of the potexvirus proteins and also indicate that potexviruses have a genome organization which is consider