Activities of SV40 T antigen necessary for the induction of tetraploid DNA content in permissive CV-1 cells

✍ Scribed by Mary Beth Albano Perry; John M. Lehman

• Publisher: John Wiley and Sons
• Year: 1998
• Tongue: English
• Weight: 231 KB
• Volume: 31
• Category: Article
• ISSN: 0196-4763
• DOI: 10.1002/(sici)1097-0320(19980401)31:4<251::aid-cyto4>3.0.co;2-m

✦ Synopsis

To determine the role of SV40 T antigen in stimulating multiple rounds of DNA synthesis in permissive cells, CV-1 cells were transfected with plasmids expressing mutant or wt T antigen in the presence or absence of the SV40 origin of replication. Induction of cells with GG2 DNA content (tetraploid DNA content) and levels of T antigen protein were detected and analyzed by flow cytometry. The mutant T antigen proteins demonstrated the expected phenotypes as determined by immunoprecipitation. Elevated levels of T antigen protein were detected in each transfection, but full-length T antigen alone was responsible for the tetraploid DNA content. The studies show that full-length T antigen with point mutations to reduce binding to the cellular proteins p53 and/or Rb were capable of inducing GG2 DNA content though the induction by these mutants was greatly enhanced by the presence of the SV40 origin of replication. Truncated T antigen (aa 1-259) could induce cells with tetraploid DNA content only in the presence of the SV40 origin of replication and the absence of Rb binidng. These studies suggest that multiple functions of T antigen are involved in the stimulation of the second round of cellular DNA synthesis.