scholarly journals Simian Virus 40 t Antigen Affects the Sensitivity of Cellular DNA Synthesis to Theophylline

1979 ◽  
Vol 30 (1) ◽  
pp. 394-396 ◽  
Author(s):  
Kathleen Rundell ◽  
Janet Cox
Keyword(s):  
Dna Synthesis ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Cellular Dna

Functional simian virus 40 T antigen is expressed in hybrid cells having finite proliferative potential

1987 ◽  
Vol 7 (4) ◽  
pp. 1541-1544
Author(s):  
O M Pereira-Smith ◽  
J R Smith
Keyword(s):  
Dna Synthesis ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Transformed Cells ◽  
Proliferative Potential ◽  
Hybrid Cells ◽  
Ras Oncogenes ◽  
Immortal Cells ◽  
Cellular Dna

Simian virus 40-transformed human cells fused with other independently derived simian virus 40-transformed cells and tumor-derived cells containing activated H-ras and N-ras oncogenes yielded hybrids capable of indefinite division. Fusions with various other immortal cells yielded hybrids that had limited division potential. T antigen expressed in limited-division hybrids was functional for the induction of cellular DNA synthesis.

scholarly journals Induction of cellular DNA synthesis by a simian virus 40 mutant defective in nuclear transport of T antigen.

1985 ◽  
Vol 5 (6) ◽  
pp. 1531-1533 ◽  
Author(s):  
R E Lanford ◽  
J K Hyland ◽  
R Baserga ◽  
J S Butel
Keyword(s):  
Dna Synthesis ◽  
Limit Of Detection ◽  
Nuclear Transport ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Quiescent Cells ◽  
Cellular Dna

The simian virus 40 (SV40) (cT)-3 mutant [SV40(cT)-3], which is defective in nuclear transport of T antigen, was utilized to determine whether cellular DNA synthesis can be stimulated by SV40 in the absence of detectable nuclear T antigen. Cellular DNA synthesis was examined in the temperature-sensitive cell cycle mutants, BHK ts13 and BHK tsAF8, after microinjection of quiescent cells with plasmid DNA containing cloned copies of wild-type SV40 or SV40(cT)-3. The efficiency of induction of cellular DNA synthesis was identical for both wild-type SV40 and SV40(cT)-3 in both cell lines. The results suggest that cell surface-associated T antigen, either alone or possibly in combination with minimal amounts of nuclear T antigen below our limit of detection, is able to stimulate cellular DNA synthesis.

Induction of cellular DNA synthesis by a simian virus 40 mutant defective in nuclear transport of T antigen

1985 ◽  
Vol 5 (6) ◽  
pp. 1531-1533
Author(s):  
R E Lanford ◽  
J K Hyland ◽  
R Baserga ◽  
J S Butel
Keyword(s):  
Dna Synthesis ◽  
Limit Of Detection ◽  
Nuclear Transport ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Quiescent Cells ◽  
Cellular Dna

The simian virus 40 (SV40) (cT)-3 mutant [SV40(cT)-3], which is defective in nuclear transport of T antigen, was utilized to determine whether cellular DNA synthesis can be stimulated by SV40 in the absence of detectable nuclear T antigen. Cellular DNA synthesis was examined in the temperature-sensitive cell cycle mutants, BHK ts13 and BHK tsAF8, after microinjection of quiescent cells with plasmid DNA containing cloned copies of wild-type SV40 or SV40(cT)-3. The efficiency of induction of cellular DNA synthesis was identical for both wild-type SV40 and SV40(cT)-3 in both cell lines. The results suggest that cell surface-associated T antigen, either alone or possibly in combination with minimal amounts of nuclear T antigen below our limit of detection, is able to stimulate cellular DNA synthesis.

Mutational analysis of simian virus 40 T antigen: stimulation of cellular DNA synthesis and activation of rRNA genes by mutants with deletions in the T-antigen gene

1983 ◽  
Vol 3 (2) ◽  
pp. 214-219
Author(s):  
K J Soprano ◽  
N Galanti ◽  
G J Jonak ◽  
S McKercher ◽  
J M Pipas ◽  
...  
Keyword(s):  
Dna Synthesis ◽  
Mutational Analysis ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Rrna Genes ◽  
Hybrid Cells ◽  
The Common ◽  
Cellular Dna ◽  
Stimulation Of

The biological activity of several deletion mutants of simian virus 40, cloned in pBR322, was determined. Three functions of the simian virus 40 A gene were studied: (i) the ability to express T antigen; (ii) the ability to induce cell DNA replication; and (iii) the ability to reactivate silent rRNA genes in hybrid cells. Recombinant plasmid DNA was introduced into cells by manual microinjection or by transfection. The results (together with previous reports) indicate that the critical sequences for these three functions are located separately on the simian virus 40 A gene, as follows: (i) the sequences necessary for the detection of the common antigenic determinant of T antigen extend from nucleotide 4147 to nucleotide 4001 (map units 0.45 to 0.42); (ii) the sequences critical for the stimulation of cell DNA synthesis extend from nucleotide 4327 to nucleotide 4001 (map units 0.49 to 0.42); and (iii) those critical for the reactivation of rRNA genes extend approximately from nucleotide 3827 to nucleotide 3526 (map units 0.39 to 0.33).

scholarly journals Functional simian virus 40 T antigen is expressed in hybrid cells having finite proliferative potential.

1987 ◽  
Vol 7 (4) ◽  
pp. 1541-1544 ◽  
Author(s):  
O M Pereira-Smith ◽  
J R Smith
Keyword(s):  
Dna Synthesis ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Transformed Cells ◽  
Proliferative Potential ◽  
Hybrid Cells ◽  
Ras Oncogenes ◽  
Immortal Cells ◽  
Cellular Dna

Simian virus 40-transformed human cells fused with other independently derived simian virus 40-transformed cells and tumor-derived cells containing activated H-ras and N-ras oncogenes yielded hybrids capable of indefinite division. Fusions with various other immortal cells yielded hybrids that had limited division potential. T antigen expressed in limited-division hybrids was functional for the induction of cellular DNA synthesis.

scholarly journals Mutational analysis of simian virus 40 T antigen: stimulation of cellular DNA synthesis and activation of rRNA genes by mutants with deletions in the T-antigen gene.

1983 ◽  
Vol 3 (2) ◽  
pp. 214-219 ◽  
Author(s):  
K J Soprano ◽  
N Galanti ◽  
G J Jonak ◽  
S McKercher ◽  
J M Pipas ◽  
...  
Keyword(s):  
Dna Synthesis ◽  
Mutational Analysis ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Rrna Genes ◽  
Hybrid Cells ◽  
The Common ◽  
Cellular Dna ◽  
Stimulation Of

The biological activity of several deletion mutants of simian virus 40, cloned in pBR322, was determined. Three functions of the simian virus 40 A gene were studied: (i) the ability to express T antigen; (ii) the ability to induce cell DNA replication; and (iii) the ability to reactivate silent rRNA genes in hybrid cells. Recombinant plasmid DNA was introduced into cells by manual microinjection or by transfection. The results (together with previous reports) indicate that the critical sequences for these three functions are located separately on the simian virus 40 A gene, as follows: (i) the sequences necessary for the detection of the common antigenic determinant of T antigen extend from nucleotide 4147 to nucleotide 4001 (map units 0.45 to 0.42); (ii) the sequences critical for the stimulation of cell DNA synthesis extend from nucleotide 4327 to nucleotide 4001 (map units 0.49 to 0.42); and (iii) those critical for the reactivation of rRNA genes extend approximately from nucleotide 3827 to nucleotide 3526 (map units 0.39 to 0.33).

Replication of cloned DNA containing the Alu family sequence during cell extract-promoting simian virus 40 DNA synthesis

1984 ◽  
Vol 4 (8) ◽  
pp. 1476-1482
Author(s):  
H Ariga
Keyword(s):  
Dna Replication ◽  
Dna Synthesis ◽  
Rna Polymerase Iii ◽  
Simian Virus 40 ◽  
Cell Extract ◽  
Simian Virus ◽  
T Antigen ◽  
Sv40 T Antigen ◽  
Sv40 Dna

The replicating activity of several cloned DNAs containing putative origin sequences was examined in a cell-free extract that absolutely depends on simian virus 40 (SV40) T antigen promoting initiation of SV40 DNA replication in vitro. Of the three DNAs containing the human Alu family sequence (BLUR8), the origin of (Saccharomyces cerevisiae plasmid 2 micron DNA (pJD29), and the yeast autonomous replicating sequence (YRp7), only BLUR8 was active as a template. Replication in a reaction mixture with BLUR8 as a template was semiconservative and not primed by a putative RNA polymerase III transcript synthesized on the Alu family sequence in vitro. Pulse-chase experiments showed that the small-sized DNA produced in a short-term incubation was converted to full-length closed circular and open circular DNAs in alkaline sucrose gradients. DNA synthesis in extracts began in a region of the Alu family sequence and was inhibited 80% by the addition of anti-T serum. Furthermore, partially purified T antigen bound the Alu family sequence in BLUR8 by the DNA-binding immunoassay. These results suggest that SV40 T antigen recognizes the Alu family sequence, similar to the origin sequence of SV40 DNA, and initiates semiconservative DNA replication in vitro.

scholarly journals Sequence-specific binding of simian virus 40 A protein to nonorigin and cellular DNA.

1984 ◽  
Vol 4 (12) ◽  
pp. 2631-2638 ◽  
Author(s):  
P J Wright ◽  
A L DeLucia ◽  
P Tegtmeyer
Keyword(s):  
Specific Binding ◽  
Consensus Sequence ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Binding Affinities ◽  
Sequence Variations ◽  
Origin Region ◽  
Cellular Dna ◽  
Many Sources

The simian virus 40 A protein (T antigen) recognized and bound to the consensus sequence 5'-GAGGC-3' in DNA from many sources. Sequence-specific binding to single pentanucleotides in randomly chosen DNA predominated over binding to nonspecific sequences. The asymmetric orientation of protein bound to nonorigin recognition sequences also resembled that of protein bound to the origin region of simian virus 40 DNA. Sequence variations in the DNA adjacent to single pentanucleotides influenced binding affinities even though methylation interference and protection studies did not reveal specific interactions outside of pentanucleotides. Thus, potential locations of A protein bound to any DNA can be predicted although the determinants of binding affinity are not yet understood. Sequence-specific binding of A protein to cellular DNA would provide a mechanism for specific alterations of host gene expression that facilitate viral function.

scholarly journals Initiation of simian virus 40 DNA replication in vitro: aphidicolin causes accumulation of early-replicating intermediates and allows determination of the initial direction of DNA synthesis.

1986 ◽  
Vol 6 (11) ◽  
pp. 3815-3825 ◽  
Author(s):  
R S Decker ◽  
M Yamaguchi ◽  
R Possenti ◽  
M L DePamphilis
Keyword(s):  
Dna Replication ◽  
Dna Synthesis ◽  
Dna Polymerase ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Early Gene ◽  
Initial Direction ◽  
Dna Polymerase Alpha

Aphidicolin, a specific inhibitor of DNA polymerase alpha, provided a novel method for distinguishing between initiation of DNA synthesis at the simian virus 40 (SV40) origin of replication (ori) and continuation of replication beyond ori. In the presence of sufficient aphidicolin to inhibit total DNA synthesis by 50%, initiation of DNA replication in SV40 chromosomes or ori-containing plasmids continued in vitro, whereas DNA synthesis in the bulk of SV40 replicative intermediate DNA (RI) that had initiated replication in vivo was rapidly inhibited. This resulted in accumulation of early RI in which most nascent DNA was localized within a 600- to 700-base-pair region centered at ori. Accumulation of early RI was observed only under conditions that permitted initiation of SV40 ori-dependent, T-antigen-dependent DNA replication and only when aphidicolin was added to the in vitro system. Increasing aphidicolin concentrations revealed that DNA synthesis in the ori region was not completely resistant to aphidicolin but simply less sensitive than DNA synthesis at forks that were farther away. Since DNA synthesized in the presence of aphidicolin was concentrated in the 300 base pairs on the early gene side of ori, we conclude that the initial direction of DNA synthesis was the same as that of early mRNA synthesis, consistent with the model proposed by Hay and DePamphilis (Cell 28:767-779, 1982). The data were also consistent with initiation of the first DNA chains in ori by CV-1 cell DNA primase-DNA polymerase alpha. Synthesis of pppA/G(pN)6-8(pdN)21-23 chains on a single-stranded DNA template by a purified preparation of this enzyme was completely resistant to aphidicolin, and further incorporation of deoxynucleotide monophosphates was inhibited. Therefore, in the presence of aphidicolin, this enzyme could initiate RNA-primed DNA synthesis at ori first in the early gene direction and then in the late gene direction, but could not continue DNA synthesis for an extended distance.

scholarly journals Simian virus 40 DNA replication in vitro: identification of multiple stages of initiation.

1989 ◽  
Vol 9 (9) ◽  
pp. 3839-3849 ◽  
Author(s):  
T Tsurimoto ◽  
M P Fairman ◽  
B Stillman
Keyword(s):  
Dna Replication ◽  
Dna Synthesis ◽  
Time Lag ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Nuclear Antigen ◽  
Replication Factor ◽  
Cellular Fraction ◽  
Template Dna

A cell-free DNA replication system dependent upon five purified cellular proteins, one crude cellular fraction, and the simian virus 40 (SV40)-encoded large tumor antigen (T antigen) initiated and completed replication of plasmids containing the SV40 origin sequence. DNA synthesis initiated at or near the origin sequence after a time lag of approximately 10 min and then proceeded bidirectionally from the origin to yield covalently closed, monomer daughter molecules. The time lag could be completely eliminated by a preincubation of SV40 ori DNA in the presence of T antigen, a eucaryotic single-stranded DNA-binding protein (replication factor A [RF-A]), and topoisomerases I and II. In contrast, if T antigen and the template DNA were incubated alone, the time lag was only partially decreased. Kinetic analyses of origin recognition by T antigen, origin unwinding, and DNA synthesis suggest that the time lag in replication was due to the formation of a complex between T antigen and DNA called the T complex, followed by formation of a second complex called the unwound complex. Formation of the unwound complex required RF-A. When origin unwinding was coupled to DNA replication by the addition of a partially purified cellular fraction (IIA), DNA synthesis initiated at the ori sequence, but the template DNA was not completely replicated. Complete DNA replication in this system required the proliferating-cell nuclear antigen and another cellular replication factor, RF-C, during the elongation stage. In a less fractionated system, another cellular fraction, SSI, was previously shown to be necessary for reconstitution of DNA replication. The SSI fraction was required in the less purified system to antagonize the inhibitory action of another cellular protein(s). This inhibitor specifically blocked the earliest stage of DNA replication, but not the later stages. The implications of these results for the mechanisms of initiation and elongation of DNA replication are discussed.

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