Temperature-Sensitive Mutants in the Study of Cell Cycle Progression in Mammalian Cells

Sheinin, Rose

doi:10.1016/b978-0-12-747750-3.50010-7

Cited by 11 publications

(5 citation statements)

References 186 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

mentioning

confidence: 99%

“…Both gene products are required for semiconservative DNA replication, which occurs normally during the DNA-synthetic, or S, phase of the cell duplication cycle [3,5,11,12,24], Because they arrest at unique stages early in S phase, they are designated DNA'7S'\ The is 2 mouse fibroblast arrests at the Gi/S interface [28] upon temperature inactivation of a polypeptide which acts to effect S-phase entry, perhaps by facilitating interaction of DNA polymerase-a and DNA primase [27]. The ts2 cell is therefore DNA'1/ Gi'1 [3,5,28]. These three ts gene products play no obligatory role in DNA repair repli cation [11,29], On the basis of these considerations and the data presented here it could be concluded that replication of mouse adenovirus DNA can occur in the absence of normal, semicon servative cellular DNA synthesis and does not require the participation of functional cellular DNA topoisomerase II orthe /.sCI or Is 2 gene products.…”

mentioning

confidence: 99%

“…Thus, the DNA of mouse adenovirus can replicate in the absence of functional DNA topoisomerase II, a DNA-chain-elongation factor, and a protein required for traverse of the Gi/S interface, respectively, encoded in the ts AIS9, /sCl and ts2 genetic loci. These results are compared with those obtained with polyoma virus.Mammalian cells temperature-sensitive (ts) in DNA synthesis (DNA'V ) were initially isolated just over one decade ago [1,2], Our interest was to use them as tools for the biochemical-genetic study of genome repli cation of, and transformation by, animal vi ruses [3][4][5][6][7]. Before such studies could be fully implemented, it was essential to accumulate a threshold of knowledge about each mutant cell, its primary functional defect, and its possible secondary expression.…”

mentioning

confidence: 99%

See 2 more Smart Citations

Mouse Polyoma Virus and Adenovirus Replication in Mouse Cells Temperature-Sensitive in DNA Synthesis

1985

Self Cite

View full text Add to dashboard Cite

Mouse adenovirus multiplies, apparently without impediment, in temperature-inactivated ts A1S9, ts C1 and ts 2 mouse fibroblasts. Thus, the DNA of mouse adenovirus can replicate in the absence of functional DNA topoisomerase II, a DNA-chain-elongation factor, and a protein required for traverse of the Gi/S interface, respectively, encoded in the ts A1S9, ts C1 and ts2 genetic loci. These results are compared with those obtained with polyoma virus.

show abstract

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 1 more Smart Citation

Mouse Polyoma Virus and Adenovirus Replication in Mouse Cells Temperature-Sensitive in DNA Synthesis

1985

Self Cite

View full text Add to dashboard Cite

show abstract

“…MATERIALS AND METHODS Cells. Wild-type (WT-4) and tsAlS9 mouse L-cells, whose properties have been described elsewhere (31)(32)(33)(34)(35)(36), were grown in suspension culture containing aminimal essential medium (40) supplemented with 7.5% (vol/vol) fetal calf serum. In all cases cells were grown to early logarithmic phase (1 x 105 to 2 x 105 cells per ml) and then mock infected or infected with virus as noted below.…”

mentioning

confidence: 99%

Synthesis of multimeric polyoma virus DNA in mouse L-cells: role of the tsA1S9 gene product

Ganz¹,

Sheinin²

1983

J Virol

View full text Add to dashboard Cite

Several different forms of progeny viral DNA can be identified in polyoma virus (Py)-infected mouse L-cells. The majority comprise mature form I superhelical DNA and the circular, double-stranded "theta" replicating intermediates in which the progeny DNA strands never exceed the unit genome length of the template. There is formed, in addition, a minority fraction of multimeric, linear, double-stranded Py DNA molecules that sediment heterogeneously at 28 to 35S and greater than 35S. Restriction enzyme analysis of these large Py DNA molecules reveals them to be tandem arrays of multiple unit genome lengths, covalently linked head to tail. It is estimated that the 28 to 35S multimeric DNA has an average size of about 20 megadaltons, made up of 6 to 20 Py genome units. The greater than 35S Py DNA is, of course, larger. Kinetic analysis indicates that formation of the monomeric progeny viral DNA and the 28 to 35S multimeric Py DNA reaches a peak at about 35 to 36 h postinfection. Synthesis of the very large linear molecules of greater than 35S is first detected after this interval and continues thereafter. The de novo synthesis of all of these progeny Py DNA molecules proceeds apparently normally in Py-infected tsA1S9 mouse L-cells incubated at 38.5 degrees C under conditions which restrict normal cellular DNA replication. These findings suggest that the cellular DNA topoisomerase II activity, encoded in the tsA1S9 locus (R. W. Colwill and R. Sheinin, submitted for publication), is not required for de novo formation of any form of Py DNA. However, the total amount made and the rate of synthesis of the large molecular weight Py DNA are affected very late in temperature-inactivated tsA1S9 cells.

show abstract

“…The ts AlS9 L cell (1) is mutant in a gene required for nuclear DNA replication (2,3) and for normal progression through the S phase of the cell cycle (4). Temperature-inactivated ts A1M9 cells synthesize "Okazald fragments" and convert them to progeny single-strand DNA of >5 x 106 daltons (2).…”

mentioning

confidence: 99%

ts A1S9 locus in mouse L cells may encode a novobiocin binding protein that is required for DNA topoisomerase II activity.

Colwill¹,

Sheinin²

1983

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Nuclear novobiocin binding proteins (NBPs) from a set of mouse L cells have been extensively purified by affinity chromatography on novobiocin-Sepharose columns. The NBPs, specifically eluted with 100 ,ug of novobiocin per ml, exhibited equivalent DNA topoisomerase activities (measured as ATP-dependent relaxation or catenation of #X174 replicative-form I DNA substrate) when extracted from equal numbers of wild-type (WT-4) mouse L cells growing logarithmically at 34WC or at 38.50C, from ts A1S9 cells similarly cultivated at the low, permissive temperature or from revertant ts+ AR cells in exponential growth at either temperature. The NBPs isolated from similar numbers of ts A1S9 cells grown to midlogarithmic phase and then incubated for 24 hr at 38.5C (the nonpermissive temperature) showed no topoisomerase II activity. Preliminary NaDodSO4/polyaerylamide gel electrophoretic analysis of enzymatically active material revealed that the NBPs of WT4 and ts+ AR cells grown at 34WC comprised three major polypeptides of 76,000, 74,000, and 30,000 daltons and a number of larger molecular mass components present in trace amounts. The NBP of ts A1S9 cells grown at the permissive temperature was similar, except that the 30,000-kilodalton polypeptide was not detected. Such enzymatically active NBPs from WT-4 and ts+ AR cells were unaffected by 100 1tg of novobiocin per ml, whereas the analogous preparation from ts A1S9 cells was totally inhibited. On the basis of these and other considerations, it is postulated that the ts A1S9 locus of mouse L cells encodes a temperature-sensitive polypeptide that is required for normal DNA topoisomerase I activity.The ts AlS9 L cell (1) is mutant in a gene required for nuclear DNA replication (2, 3) and for normal progression through the S phase of the cell cycle (4). Temperature-inactivated ts A1M9 cells synthesize "Okazald fragments" and convert them to progeny single-strand DNA of >5 x 106 daltons (2). In addition they support full replication of monomeric and multimeric (up to 34 X 107 daltons) polyoma DNA (5, 6). Therefore, it was concluded that the enzyme proteins of polydeoxyribonucleotide guished by their properties and by the mechanism by which they modify the topology of DNA. The DNA topoisomerase I more closely resembles the nicking-closing, w protein activity already well characterized biochemically and genetically in bacteria (12, 13). Whereas the topoisomerase I enzyme has long been recognized to function in eukaryotes (14), evidence for the topoisomerase II eukaryotic analogue for the DNA gyrase of prokaryotes was reported later (15-17) and, indeed, after the studies described herein were initiated. No evidence for abnormal topoisomerase I activity had been seen in earlier studies of temperature-inactivated ts AlS9 cells (cf. refs. 4 and 6; unpublished data). Therefore, attention was focused on DNA topoisomerase II. The first hint that this enzyme might be affected came with the demonstration that ts AlS9 cells are hypersensitive to novobiocin (9) The publication co...

show abstract

Temperature-Sensitive Mutants in the Study of Cell Cycle Progression in Mammalian Cells

Cited by 11 publications

References 186 publications

Mouse Polyoma Virus and Adenovirus Replication in Mouse Cells Temperature-Sensitive in DNA Synthesis

Mouse Polyoma Virus and Adenovirus Replication in Mouse Cells Temperature-Sensitive in DNA Synthesis

Synthesis of multimeric polyoma virus DNA in mouse L-cells: role of the tsA1S9 gene product

ts A1S9 locus in mouse L cells may encode a novobiocin binding protein that is required for DNA topoisomerase II activity.

Contact Info

Product

Resources

About