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Replicative functions of the SV40(cT)-3 mutant defective for nuclear transport of T antigen
Abstract:
The SV40(cT)-3 mutant is defective in transport of SV40 large tumor antigen (T-ag) to the nucleus. Several properties of T-ag associated with SV40 lytic infection and attributed to its nuclear localization were examined to determine whether biologically significant levels of the mutant T-ag (cT-ag) that were immunologically undetectable were transported to the nucleus in SV40(cT)-3-infected TC-7 cells. SV40(cT)-3 was defective in regulation of T-ag synthesis and initiation of viral DNA synthesis. These defects were presumably due to the lack of nuclear transport of cT-ag, since cT-ag was capable of interacting with the SV40 origin of viral DNA synthesis in a solution binding assay. The level of fatty acid acylation, a modification specific for the cell surface associated T-ag, was not affected by the cT mutation. The cT mutation sufficiently suppressed the nuclear transport of wild-type (WT) T-ag in SV40(cT)-3-infected COS-1 cells to result in the cessation of WT-T-ag-stimulated SV40(cT)-3 viral DNA synthesis. These results are discussed with respect to the recent findings that SV40(cT)-3 is fully competent for the transformation of established cell lines and the induction of cellular DNA synthesis in quiescent cells.
Insights
The SV40(cT)-3 mutant impairs nuclear transport of large tumor antigen (T-ag), affecting viral DNA synthesis regulation. Despite this defect, the mutant retains cell transformation and DNA synthesis induction capabilities.
Area of Science:
- Molecular Biology
- Virology
- Cell Biology
Background:
- Simian virus 40 (SV40) large tumor antigen (T-ag) plays crucial roles in viral lytic infection.
- Nuclear localization of T-ag is essential for its functions, including viral DNA synthesis regulation.
Purpose of the Study:
- To investigate the nuclear transport of the SV40(cT)-3 mutant T-ag (cT-ag).
- To determine if biologically significant levels of cT-ag reach the nucleus.
- To assess the impact of impaired nuclear transport on viral DNA synthesis and T-ag functions.
Main Methods:
- Analysis of T-ag nuclear transport in infected TC-7 and COS-1 cells.
- Examination of T-ag synthesis regulation and viral DNA synthesis initiation.
- Solution binding assay to assess cT-ag interaction with the SV40 origin of DNA synthesis.
- Evaluation of fatty acid acylation levels on T-ag.
Main Results:
- SV40(cT)-3 mutant is defective in T-ag nuclear transport.
- Defects in T-ag synthesis regulation and viral DNA synthesis initiation were observed.
- cT-ag demonstrated interaction with the SV40 origin of DNA synthesis in vitro.
- The cT mutation suppressed wild-type T-ag nuclear transport, halting viral DNA synthesis.
Conclusions:
- Impaired nuclear transport of cT-ag is responsible for defects in SV40 lytic cycle regulation.
- Despite nuclear transport defects, SV40(cT)-3 remains competent for cell transformation and induction of cellular DNA synthesis.