ANALYSIS OF TS2, A TEMPERATURE-SENSITIVE MUTANT IN DNA SYNTHESIS.
Item
-
Title
-
ANALYSIS OF TS2, A TEMPERATURE-SENSITIVE MUTANT IN DNA SYNTHESIS.
-
Identifier
-
AAI8515645
-
identifier
-
8515645
-
Creator
-
MALKAS, LINDA HELEN.
-
Contributor
-
Harvey L. Ozer
-
Date
-
1985
-
Language
-
English
-
Publisher
-
City University of New York.
-
Subject
-
Chemistry, Biochemistry
-
Abstract
-
A temperature-sensitive mutant (ts2) of the mouse cell line Balb/3T3 affected in DNA synthesis was further characterized. FMF analysis of actively growing ts2 cells shifted to the non-permissive temperature, 39(DEGREES)C, suggested an accumulation of these cells within the S phase with increasing incubation time at 39(DEGREES)C.;Serum stimulation of synchronized ts2 cells showed an induction of DNA synthesis and the enzymes thymidine kinase, thymidylate kinase, and DNA polymerase (alpha) at 33(DEGREES)C, while exhibiting little (or no) stimulation of these activities in cells incubated at 39(DEGREES)C. Similar experiments performed with the ts2 parent cell line and a revertant showed the stimulation of these activities at both 33(DEGREES)C and 39(DEGREES)C.;These synchronized ts2 cells did not show an increased requirement for serum at 39(DEGREES)C, and were fully capable of entering the cell cycle at 39(DEGREES)C; as indicated by an increase in cellular protein content, and the induction of both c-myc and proliferin mRNA's during the time course. The induction of enzyme activity was not dependent on the stimulation of DNA synthesis, since TK was induced in the presence of hydroxyurea. These results suggest that the lack of enzyme induction in growth-arrested ts2 cells at 39(DEGREES)C after the addition of serum is associated with a broad cellular phenomena; the result of the ts lesion.;SV40 infection of quiescent ts2 cells stimulated the rate of ('3)H-TdR incorporation and TK activity in these cells at both 33(DEGREES)C and 39(DEGREES)C. The induction of DNA synthesis was shown to be dependent on the presence of the SV40 large T-antigen. Those cells stimulated into the cell cycle by SV40 infection were able to complete DNA replication and progress through one round of cell division at 33(DEGREES)C and 39(DEGREES)C. SV40 transformants of ts2 were isolated to assess whether the continual presence of the T-antigen would complement the ts mutation. These transformants were found to be unable to support cell growth at 39(DEGREES)C.;A model postulated to explain the observed aberrant response of ts2 cells at 39(DEGREES)C considers a broadly based transcriptional block at the non-permissive temperature.
-
Type
-
dissertation
-
Source
-
PQT Legacy CUNY.xlsx
-
degree
-
Ph.D.
-
Program
-
Biochemistry
