Characterization of the interaction of polyomavirus middle T antigen with type 2A protein phosphatase.
Two cellular proteins of 36 and 63 kDa which bind the small T and middle T antigens of polyomavirus recently have been identified as the catalytic and regulatory subunits of the phosphoserine/threonine-specific type 2A protein phosphatase (PP2A). We report here the presence of phosphoseryl phosphatase activity associated with polyomavirus small T and middle T antigens in immunoprecipitates prepared from virus-infected and transformed cells. Phosphatase activity was also found associated with middle T-antigen mutants, some of which had been defined previously to associate with 36- and 63-kDa cellular proteins. Middle T-antigen-associated phosphatase activity was sensitive to okadaic acid and microcystin-LR, inhibitors of PP2A, and insensitive to inhibitor 1 or 2, orthovanadate, or EDTA. Using antiserum specific for the catalytic subunit of PP2A, we found that unlike the majority of PP2A, middle T-antigen-bound PP2A was membrane associated. However, no gross change in the amount, activity, or localization of PP2A could be attributed to middle T-antigen expression in transformed cells. Anti-PP2A antibodies coprecipitated a 63-kDa protein from normal cells and in addition coprecipitated middle T antigen, 60- and 61-kDa proteins (identified as src family members), and an 81-kDa protein from middle T-antigen-transformed cells. Furthermore, we detected protein kinase activity in PP2A immunoprecipitates and protein phosphatase activity in src immune complexes from extracts of middle T-antigen-transformed, but not normal, cells. These results reinforce the notion that at least a portion of middle T antigen bridges a protein kinase with a protein phosphatase.
Ulug
E T
ET
Section of Virology and Oncology, Kansas State University, Manhattan 66506-4901.
Cartwright
A J
AJ
Courtneidge
S A
SA
D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000378:metabolism*; D002462:Cell Membrane / Q000201:enzymology; D002478:Cells, Cultured; D046911:Macromolecular Substances; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010768:Phosphoserine / Q000378:metabolism; D011485:Protein Binding; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism; D051381:Rats
D000952:Antigens, Polyomavirus Transforming; D046911:Macromolecular Substances; D010768:Phosphoserine; D011505:Protein-Tyrosine Kinases; D016392:Proto-Oncogene Proteins pp60(c-src); D010749:Phosphoprotein Phosphatases