Lytic functions of mutant polyomavirus large T-antigen with deletion of retinoblastoma protein-binding motif.
Polyomavirus large T-antigen has a binding site for the retinoblastoma protein. Mutant proteins without this binding site retain their activity in the initiation of viral DNA synthesis, but are not able to induce immortalization of rat embryo fibroblasts. To investigate the function of the retinoblastoma binding site in polyomavirus lytic infection, a set of mutants with small in-frame deletions was constructed. We found that the mutant large T-antigens had partial or complete defects in immortalization and had corresponding disabilities in trans-activation of the adenovirus E2 promoter in secondary rat embryo cells. However, one partially immortalization-defective mutant trans-activated the E2 promoter in HeLa cells. All mutants were normal in trans-activation of the polyomavirus late promoter. The mutant large T-antigens had a decreased activity in viral DNA replication. This phenotype was observed only after transfection of growth-arrested mouse cells. The replication of the mutants could be restored to nearly wild-type levels when middle and small T-antigens were coexpressed in the cells. The same effect was achieved with a 287-amino-acid N-terminal fragment of large T-antigen with an intact retinoblastoma protein-binding motif. This result suggests that the effect on viral DNA synthesis was indirect.
Department of Medical Virology, Uppsala University Biomedical Centre, Sweden.
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Söderbärg
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Magnusson
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000235:genetics / Q000502:physiology*; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics / Q000502:physiology*; D002478:Cells, Cultured; D017353:Gene Deletion; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D011120:Polyomavirus / Q000235:genetics / Q000502:physiology; D051381:Rats; D016160:Retinoblastoma Protein / Q000378:metabolism*; D015533:Transcriptional Activation / Q000502:physiology; D014779:Virus Replication / Q000235:genetics / Q000502:physiology*
D000952:Antigens, Polyomavirus Transforming; D002352:Carrier Proteins; D016160:Retinoblastoma Protein