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A new SV40 mutant that encodes a small fragment of T antigen transforms established rat and mouse cells
1Department of Molecular, Cellular, and Developmental Biology, University of Colorado, Boulder 80309.
Abstract:
We have constructed a new SV40 mutant, T147, that makes a large T antigen that is only 147 amino acids long. We show that the T147 T antigen is a phosphoprotein that is as stable as wild-type T antigen and that the SV40 viral origin binding activity of the T147 T antigen is reduced at least 100-fold relative to that of wild-type T antigen. Most importantly, we demonstrate that cloned T147 DNA transforms rat F111 cells to anchorage independence as efficiently as cloned wild-type SV40 DNA and that cloned T147 DNA also efficiently transforms C3H10T1/2 mouse cells in a focus assay.
Insights
A new Simian virus 40 (SV40) mutant, T147, produces a truncated large T antigen. Despite reduced viral origin binding, this mutant efficiently transforms cells, indicating its oncogenic potential.
Area of Science:
- Molecular Biology
- Virology
- Cancer Research
Background:
- Simian virus 40 (SV40) large T antigen is a key viral oncoprotein.
- Understanding the structure-function relationship of large T antigen is crucial for cancer research.
Purpose of the Study:
- To investigate the properties and transforming activity of a novel, truncated SV40 large T antigen (T147).
- To assess the impact of a 147-amino acid truncation on T antigen stability, DNA binding, and cellular transformation.
Main Methods:
- Construction and characterization of the SV40 T147 mutant.
- Biochemical analysis of T147 T antigen stability and phosphoprotein status.
- Assessment of SV40 viral origin binding activity.
- Cellular transformation assays using rat F111 and mouse C3H10T1/2 cells.
Main Results:
- The T147 T antigen is a stable phosphoprotein, comparable to wild-type.
- SV40 viral origin binding activity of T147 T antigen is reduced over 100-fold.
- Cloned T147 DNA efficiently transforms rat F111 cells to anchorage independence.
- Cloned T147 DNA also efficiently transforms mouse C3H10T1/2 cells in a focus assay.
Conclusions:
- Truncated SV40 T147 T antigen retains significant cellular transforming capabilities despite impaired DNA binding.
- The study highlights that regions outside the DNA-binding domain may be critical for SV40-mediated transformation.
- T147 represents a valuable tool for dissecting the oncogenic mechanisms of SV40 T antigen.