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1.
Recurring defective variants of simian virus 40 containing monkey DNA segments. 总被引:4,自引:2,他引:2
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Four independently and newly isolated defective variants of simian virus 40 have been characterized. All four are very similar, if not identical, to two previously and independently isolated variants (Wakamiya et al., J. Biol. Chem. 254:3584-3591, 1979; J. Papamatheakis, E. Kuff, E. Winocour, and M. F. Singer, J. Biol. Chem. 255:8919-8927, 1980). The documented similarities include restriction endonuclease maps and the presence of the same monkey DNA segments covalently linked to simian virus 40 DNA sequences. Each of the newly described variants was first detected upon serial passaging of wild-type simian virus 40 at a high multiplicity of infection at 33 degrees C as recently described (M. F. Singer and R. E. Thayer, J. Virol. 35:141-149, 1980). A variety of experiments support the idea that the various isolates were independent and do not reflect inadvertent cross-contamination. Two of the new isolates arose during passage of wild-type strain 777 virus in BSC-1 cells, one during passage of strain 776 in BSC-1 cells, and one during passage of strain 776 in primary African green monkey kidney cells. The two variants obtained after passage of strain 776 were shown to contain a particular recognition site for restriction endonuclease MboII within their simian virus 40 DNA segments, as do the two previous isolates. This site is not present in wild-type strain 776 DNA but is shown here to be present in wild-type strain 777 DNA. The surprising recurrence of closely related variants and particularly the unexpected presence of the endo R.MboII site in variants derived from passaging strain 776 suggest that these variants may arise by mechanisms other than recombination between the initial infecting viral genome and the host DNA. 相似文献
2.
Structure of a newly isolated variant of Simian virus 40 DNA containing monkey DNA and its similarity to previously isolated variants 总被引:1,自引:0,他引:1
J D Papamatheakis E L Kuff E Winocour M Singer 《The Journal of biological chemistry》1980,255(18):8919-8927
The structure of a newly and independently isolated defective variant of simian virus 40 that contains covalently linked monkey and SV40 DNA sequences is described. This variant, termed 290, has a structure essentially identical with a previously isolated and characterized variant named CVP8/1/P2 (Eco RI res). The structural similarities include the monkey (host) DNA segment that is combined with viral DNA sequences, the particular viral DNA segment that is present, and the arrangement of these within the defective genome. The monkey DNA segment contains sequences derived from both low and high reiteration frequency monkey DNA. The viral sequences include the origin of replication. The separate isolation of essentially identical variants suggests a high level of specificity in the events leading to the formation and amplification of this type of defective genome. 相似文献
3.
In an effort to characterize sites of recombination between SV40 and monkey DNA, we have determined the primary sequence of a large portion of the SV40 variant, designated 1103. This virus contains DNA sequences derived both from the wild type SV40 genome and from the permissive monkey cell in which the virus was propagated. Further, the monkey sequences included in the defective genome are homologous to both highly repeated monkey DNA (alpha component) and sequences that are infrequently repeated in the monkey genome. The regions of the 1103 genome where DNA sequences were determined include 1) the segments of the variant that surround joints connecting SV40 and monkey sequences, 2) the segment that contains the joint between monkey sequences of high and low reiteration frequency, and 3) the DNA segment of the variant that is homologous to monkey alpha component DNA. Comparison of the data obtained from the sequences analysis of the SV40 variants 1103 and CVP8/1/P2 (EcoRI res) (described in Wakamiya, T., McCutchan, T., Rosenberg, M., and Singer, M. (1979) J. Biol. Chem 254, 3584-3591) reveals certain similarities between the two that may be involved in eukaryotic recombination and defective variant formation. 相似文献
4.
Isolation and characterization of defective simian virus 40 genomes which complement for infectivity. 总被引:1,自引:1,他引:0
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A new variant of simian virus 40 (EL SV40), containing the complete viral DNA separated into two molecules, was isolated. One DNA species contains nearly all of the early (E) SV40 sequences, and the other DNA contains nearly all of the late (L) viral sequences. Each genome was encircled by reiterated viral origins and termini and migrated in agarose gels as covalently closed supercoiled circles. EL SV40 or its progenitor appears to have been generated in human A172 glioblastoma cells, as defective interfering genomes during acute lytic infections, but was selected during the establishment of persistently infected (PI) green monkey cells (TC-7). PI TC-7/SV40 cells contained EL SV40 as the predominant SV40 species. EL SV40 propagated efficiently and rapidly in BSC-1, another line of green monkey cells, where it also formed plaques. EL SV40 stocks generated in BSC-1 cells were shown to be free of wild-type SV40 by a number of criteria. E and L SV40 genomes were also cloned in the bacterial plasmid pBR322. When transfected into BSC-1 cell monolayers, only the combination of E and L genomes produced a lytic infection, followed by the synthesis of EL SV40. However, transfection with E SV40 DNA alone did produce T-antigen, although at reduced frequency. 相似文献
5.
Acquisition of Sequences Homologous to Host DNA by Closed Circular Simian Virus 40 DNA II. Further Studies on the Serial Passage of Virus Clones 总被引:8,自引:5,他引:3
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Sara Lavi Shmuel Rozenblatt Maxine F. Singer Ernest Winocour 《Journal of virology》1973,12(3):492-500
Three plaque isolates of SV40 strain 777 and 1 plaque isolate of strain 776 were grown to high-titer stocks and serially passaged, undiluted, in monkey BS-C-1 cells. In each case, the serial passaging procedure resulted in the accumulation of closed-circular SV40 DNA molecules containing covalently linked sequences homologous to reiterated host cell DNA (called substituted virus DNA). The relative yields, at a given passage level, of SV40 DNA with measurable homology to host DNA varied in different sets of serial passages, including passages of the same virus clone. More reproducible yields of substituted viral DNA progeny were obtained when the serial passaging procedure was initiated from earlier passages rather than from the original plaque-purified stock. Fractionation of closed-circular SV40 DNA molecules on alkaline sucrose gadients indicated that the majority of substituted virus DNA molecules are not plaque producers and are slightly smaller in size than plaque-forming DNA molecules which display no detectable homology to host DNA. Evidence that substituted SV40 DNA molecules replicate during serial undiluted passage was obtained from experiments which demonstrated (i) the presence of host sequences in replicative forms of the viral DNA and (ii) the incorporation of (3)H-thymidine into host sequences isolated from the mature substituted virus DNA molecule. 相似文献
6.
The adenovirus type 2-simian virus 40 hybrid virus Ad2+ND4 requires deletion variants to grow in monkey cells. 总被引:2,自引:2,他引:0
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The Ad2+ND4 virus is an adenovirus type 2 (Ad2)-simian virus 40 (SV40) recombination. The Ad2 genome of this recombinant has a rearrangement within early region 3; Ad2 DNA sequences between map positions 81.3 and 85.5 have been deleted, and the SV40 DNA sequences between map positions 0.11 and 0.626 have been inserted into the deletion in an 81.3-0.626 orientation. Nonhybrid Ad2 is defective in monkey cells; however, the Ad2+ND4 virus can replicate in monkey cells due to the expression of the SV40-enhancing function encoded by the DNA insert. Stocks of the Ad2+ND4 hybrid were produced in primary monkey cells by using the progeny of a three-step plaque purification procedure and were considered to be homogeneous populations of Ad2+ND4 virions because they induced plaques in primary monkey cells by first-order kinetics. By studying the kinetics of plaque induction in continuous lines (BSC-1 and CV-1) of monkey cells, we have found that stocks (prepared with virions before and after plaque purification) of Ad2+ND4 are actually heterogeneous populations of Ad2+ND4 virions and Ad2+ND4 deletion variants that lack SV40 and frequently Ad2 DNA sequences at the left Ad2-SV40 junction. Due to the defectiveness of the Ad2+ND4 virus, the production of progeny in BSC-1 and CV-1 cells requires complementation between the Ad2+ND4 genome and the genome of an Ad2+ND4 deletion variant. Since the deletion variants that have been obtained from Ad2+ND4 stocks do not express the SV40-enhancing function in that they cannot produce progeny in monkey cells, we conclude that they are providing an Ad2 component that is essential for the production of Ad2+ND4 progeny. These data imply that the Ad2+ND4 virus is incapable of replicating in singly infected primary monkey cells without generating deletion variants that are missing various amounts of DNA around the left Ad2-SV40 junction in the hybrid genome. As the deletion variants that arise from the Ad2+ND4 virus are created by nonhomologous DNA recombination, the generation of deletion variants in monkey cells infected with Ad2+ND4 may be a useful model for studying this process. 相似文献
7.
Deoxyribonucleic Acid Replication in Simian Virus 40-Infected Cells: III. Comparison of Simian Virus 40 Lytic Infection in Three Different Monkey Kidney Cell Lines
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A comparative study of simian virus 40 (SV40) lytic infection in three different monkey cell lines is described. The results demonstrate that viral deoxyribonucleic acid (DNA) synthesis and infectious virus production begin some 10 to 20 hr earlier in CV-1 cells and primary African green monkey kidney (AGMK) cells than in BSC-1 cells. Induction of cellular DNA synthesis by SV40 was observed in CV-1 and AGMK cells but not with BSC-1 cells. Excision of large molecular weight cellular DNA to smaller fragments was easily detectable late in infection of AGMK cells. Little or no excision was observed at comparable times after infection of CV-1 and BSC-1 cells. The different kinds of responses of these three monkey cell lines during SV40 lytic infection suggest the involvement of cellular functions in the virus-directed induction of cellular DNA synthesis and the excision of this DNA from the genome. 相似文献
8.
The growth of simian virus 40 (SV40) host range/adenovirus helper function mutants in an African green monkey cell line that constitutively expresses the SV40 agnoprotein. 总被引:1,自引:1,他引:0
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The simian virus 40 T-antigen carboxy-terminal mutants, dlA2459 and dlA2475, are cell line and temperature dependent for growth and plaque formation in monkey kidney cells. Although these mutants did form plaques on BSC-1 cells at 37 degrees C, they were about fivefold less efficient for plaque formation than wild-type simian virus 40. These mutants did not grow in CV-1 cells and did not synthesize agnoprotein in those cells. CV-1 cells which constitutively express the agnoprotein were permissive for mutant plaque formation. However, late mRNAs, virion proteins, and progeny virion yields did not accumulate to wild-type levels during mutant infection of the agnoprotein-producing cells. 相似文献
9.
Incorporation of Iododeoxyuridine into Cellular Deoxyribonucleic Acid Synthesized After Infection with Simian Virus 40 总被引:1,自引:0,他引:1
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Primary monkey kidney cells (Cerocpithecus aethiops) in the stationary phase of growth were labeled with (14)C-thymidine for 24 hr prior to infection with simian virus 40 (strain 777). (3)H-deoxyadenosine and 5-iodo-2'-deoxyuridine (IUdR) were added to some of the cultures 24, 48, or 72 hr after infection; 24 hr later the deoxyribonucleic acid (DNA) was extracted from these cultures and centrifuged in a CsCl density gradient. The portion of DNA which had become heavier because of incorporation of IUdR could be seen as a second peak in the sedimentation profile. This peak contained (14)C as well as (3)H activity. The possibility that the (14)C-labeled cellular DNA might be degraded and used for the synthesis of viral DNA could be excluded. On the basis of these results, it must be assumed that the infection of monkey kidney cells with simian virus 40 induces the synthesis of cellular DNA. 相似文献
10.
Chromatin structure of simian virus 40-pBR322 recombinant plasmids in COS-1 cells. 总被引:7,自引:2,他引:5
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To study the nucleoprotein structure formed by recombinant plasmid DNA in mammalian cells, nuclei were isolated from COS-1 cells after transfection with a recombinant (pJI1) containing pBR322 sequences and a segment of simian virus 40 containing information for a nuclease-sensitive chromatin structure. The nuclei were incubated with DNase I. DNA fragments which were the size of linear pJI1 DNA were isolated, redigested with restriction enzymes, fractionated by electrophoresis, and detected by hybridization with nick-translated segments prepared from the plasmid DNA. Two DNase I-sensitive sites were detected in the simian virus 40 portion of the plasmid at the same sites that were DNase I sensitive in simian virus 40 chromatin prepared late after infection of African green monkey kidney (BSC-1) cells. One site extended from the viral origin of replication to approximately nucleotide 40. The 21-base pair repeated sequences were relatively DNase I resistant. A second site occurred over the single copy of the 72-base pair segment present in this plasmid. These results indicate that the nuclease-sensitive chromatin structure does not depend on the presence of viral structural proteins. In addition, late viral proteins added to pJI1-transfected COS-1 cells by superinfection with simian virus 40 caused no change in the distribution of DNase I-sensitive sites in plasmid chromatin. Analysis of transfected plasmid DNA may provide a general method applicable to the study of the chromatin structure of cloned segments of DNA. 相似文献
11.
Mutations near the carboxyl terminus of the simian virus 40 large tumor antigen alter viral host range. 总被引:23,自引:17,他引:6
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J M Pipas 《Journal of virology》1985,54(2):569-575
We report the characterization of three mutants of simian virus 40 with mutations that delete sequences near the 3' end of the gene encoding large tumor antigen (T antigen). Two of these mutants, dl1066 and dl1140, exhibit an altered viral host range. Wild-type simian virus 40 is capable of undergoing a complete productive infection on several types of established African green monkey kidney lines, including BSC40 and CV1P. dl1066 and dl1140 grow on BSC40 cells at 37 degrees C. However, both mutants fail to form plaques on BSC40 cells at 32 degrees C or on CV1P cells at any temperature. These mutants are capable of replicating viral DNA in the nonpermissive cell type, indicating a defect in an activity of T antigen not related to its replication function. Furthermore this defect can be complemented in trans by the wild type or by a variety of DNA replication-negative T antigen mutants, so long as they produce a normal carboxyl-terminal region of the molecule. Our data are consistent with the hypothesis that the C-terminal region of T antigen constitutes a functional domain. We propose that this domain encodes an activity that is required for simian virus 40 productive infection on the CV1P cell line, but not on BSC40. 相似文献
12.
13.
Recombination in simian virus 40-infected cells. Structure of naturally arising variants ev-2114, ev-2102, and ev-1110 总被引:6,自引:0,他引:6
The complete nucleotide sequence has been determined for three newly cloned evolutionary variants from two different independently generated evolutionary series (1100 and 2100 series) of simian virus 40 (SV40). These naturally arising variants, designated ev-1110, ev-2102, and ev-2114, were isolated after five high multiplicity serial passages. The structure of the variants consists of a monomeric unit tandemly repeated four times (ev-2102 and ev-2114) or six times (ev-1110) in the variant genome; the variants have four or six copies, respectively, of the viral origin signal for DNA replication. The DNA content in the three variants is vastly different in that the genome of variant ev-2114 contains only rearranged viral sequences, while variant ev-2102 contains a substitution with monkey DNA sequences consisting of a nearly complete dimeric unit of Alu family sequences as well as less repetitive sequences and variant ev-1110 contains monkey DNA sequences derived solely from repetitive alpha-component DNA. Recombination events, cellular sequences, and structural features of these and other naturally arising SV40 variants are compared. 相似文献
14.
T Wakamiya T McCutchan M Rosenberg M Singer 《The Journal of biological chemistry》1979,254(9):3584-3591
The entire nucleotide sequence (1210-base-pair repeating units) of a defective variant of simian virus 40 is presented. Within this variant there are deletions of large portions of the wild type genome and an inversion within the remaining wild type viral sequences. In addition, the defective variant contains DNA sequences derived from the permissive monkey cells in which the virus was propagated. The monkey sequences include a portion that is homologous to sequences within highly repeated monkey DNA (alpha component) as well as portions derived from sequences that are infrequently repeated in the monkey genome. One out of every three to four of the tandem 1210-base-pair repeat units contains in addition, a duplication of a part of the monkey sequences. The sequence information defines the structures of a number of recombinational joints which result from deletions, inversions, duplications, and insertions of host sequences into the viral genome. The data demonstrate that the various recombinational events that resulted in the formation of this defective variant did not depend on extensive homology between recombining segments. 相似文献
15.
16.
Genomic arrangement of an adenovirus-simian virus 40 hybrid virus, Ad2+ND4del. 总被引:8,自引:6,他引:2
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H Westphal 《Journal of virology》1981,40(2):526-532
Ad2+ND4del is an adenovirus type 2-simian virus 40 hybrid virus nondefective for growth in human cells. The virus was first observed when stocks of Ad2+ND4, a hybrid isolated from primary monkey kidney cells, were propagated in human cells. This paper describes the DNA sequence at two sites of DNA recombination, the site of the left adenovirus type 2-simian virus 40 junction and the site of a deletion of internal simian virus 40 sequences. Since the deletion was observed when the virus was switched from monkey to human cells, an analysis of gene expression in the region of DNA rearrangement may prove useful for the elucidation of molecular events that accompany virus growth in different hosts. 相似文献
17.
Expression of Epstein-Barr viral early antigen in monolayer tissue cultures after transfection with viral DNA and DNA fragments. 总被引:10,自引:3,他引:7
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Antigens associated with the Epstein-Barr virus (EBV) replicative cycle were found in the nucleus and cytoplasm of human placental, Vero, BSC-1, and owl monkey kidney cells transfected with EBV DNA prepared from several different strains of virus. The number of antigen-positive nuclei increased when transfection was followed by cell fusion induced by inactivated Sendai virus. About 1,200 antigen-positive foci were induced per micrograms of EBV DNA. On the basis of their reactivity with various well-characterized human sera, it appears that the antigens are part of the early antigen complex. None of the four restriction endonucleases, EcoRI, HindIII, SalI, and BamHI, destroyed the ability of EBV DNA to induce early antigen. However, only SalI seemed to leave intact the full spectrum of antigen expression by the HR-1 and FF41 strains of EBV DNA. By means of transfection with recombinant DNA plasmids containing different EBV (FF41) DNA fragments regenerated by EcoRI, we showed that the coding region for early antigen was at least partially contained on the 17.2-megadalton EcoRI B fragment. 相似文献
18.
Properties of permissive monkey cells transformed by UV-irradiated simian virus 40. 总被引:7,自引:13,他引:7
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African green monkey cells (CV1 line) were infected with UV-irradiated simian virus 40 (SV40), and permissive lines of stably transformed cells were established. These cell lines display the SV40 T-antigen and the growth characteristics typical of nonpermissive transformed cells (e.g., reduced cell density inhibition, reduced serum dependence, ability to overgrow normal cells, and colony formation in soft agar). The level of permissiveness to superinfecting SV40 is fully comparable with that of nontransformed CV1 and BSC-1 lines. The transformed monkey lines also support SV40 plaque production under agar. By Cot analysis, the transformed permissive cells contain, on an average, 1 to 2 SV40 genome equivalents, and the majority of the viral sequences are associated with the high-molecular-weight cellular DNA. No spontaneous production of infectious SV40 has been observed. The transformed permissive monkey cells failed to support the replication of SV40 tsA mutants at the restrictive temperature. To account for this, it is suggested that the gene A product has separate functions for transformation and initiation of viral DNA synthesis, and only the former function is expressed in the transformed permissive monkey cells. 相似文献
19.
Replication of simian virus 40 DNA in normal human fibroblasts and in fibroblasts from xeroderma pigmentosum. 总被引:8,自引:1,他引:7
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Simian virus 40 infection of semipermissive human diploid fibroblasts (HF), at early passage in cell culture, was compared with that of permissive established monkey cell lines. Viral DNA can be readily detected at 24 to 48 h postinfection at 37 degrees C with a high multiplicity of infection, approaching 10% of that of monkey cells (TC7). The length of time necessary for replication of an average molecule of viral DNA was found to be indistinguishable in HF and TC7 cells. Strand elongation plus termination were assessed by following the accumulation of DNA I at 40 degrees C from replicative intermediates of tsA30 prelabeled at 33 degrees C, obviating isotope pool problems. Combined initiation and elongation of wild-type viral DNA was measured by density shift experiments involving a 5-bromodeoxyuridine chase of prelabeled [3H]thymidine-labeled viral DNA. Determination of accumulation of viral T and V antigens supports the conclusion that the most likely basis for the reduced virus yield in HF cells results from the inefficiency of an early stage in virus infection, before or during uncoating. Similar results were obtained in fibroblasts derived from patients with xeroderma pigmentosum, suggesting that enzymes of UV repair are not required in unirradiated simian virus 40 DNA synthesis. 相似文献
20.
Post-transcriptional restriction of human adenovirus expression in monkey cells. 总被引:17,自引:17,他引:0
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L Eron 《Journal of virology》1975,15(5):1256-1261
Infection of the continuous simian cell lines CV-1 and BSC-1 with human adenovirus type 2 (Ad2) is abortive. However, the restriction of Ad2 reproduction in these cells can be overcome by increasing the Ad2 infectious dose or by coinfection with simian virus 40. Vero, another established simian cell line free of detectable endogenous simian virus 40 DNA, is not restricted in its ability to promote Ad2 growth even at low input multiplicities of Ad2 and in the absence of SV40 helper. The amount of structural Ad2 proteins in total cell extracts of enhanced BSC-1 cells is at least two orders of magnitude higher than that of unenhanced cells. In contrast, comparable quantities of Ad2 mRNA specifying these proteins are found in both the enhanced and the unenhanced cell. Both sets of mRNA can be translated in a cell-free system with equal efficiency. 相似文献