Catalytically inactive protein phosphatase 2A can bind to polyomavirus middle tumor antigen and support complex formation with pp60(c-src).
Interaction between the heterodimeric form of protein phosphatase 2A (PP2A) and polyomavirus middle T antigen (MT) is required for the subsequent assembly of a transformation-competent MT complex. To investigate the role of PP2A catalytic activity in MT complex formation, we undertook a mutational analysis of the PP2A 36-kDa catalytic C subunit. Several residues likely to be involved in the dephosphorylation mechanism were identified and mutated. The resultant catalytically inactive C subunit mutants were then analyzed for their ability to associate with a cellular (B subunit) or a viral (MT) B-type subunit. Strikingly, while all of the inactive mutants were severely impaired in their interaction with B subunit, most of these mutants formed complexes with polyomavirus MT. These findings indicate a potential role for these catalytically important residues in complex formation with cellular B subunit, but not in complex formation with MT. Transformation-competent MT is known to associate with, and modulate the activity of, several cellular proteins, including pp60(c-src) family kinases. To determine whether association of MT with an active PP2A A-C heterodimer is necessary for subsequent association with pp60(c-src), catalytically inactive C subunits were examined for their ability to form complexes containing pp60(c-src) in MT-expressing cells. Two catalytically inactive C subunit mutants that efficiently formed complexes with MT also formed complexes that included an active pp60(c-src) kinase, demonstrating that PP2A activity is not essential in cis in MT complexes for subsequent pp60(c-src) association.
Ogris
E
E
Division of Cellular and Molecular Biology, Dana-Farber Cancer Institute, Department of Pathology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Mudrak
I
I
Mak
E
E
Gibson
D
D
Pallas
D C
DC
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000378:metabolism*; D002384:Catalysis; D020134:Catalytic Domain; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010749:Phosphoprotein Phosphatases / Q000235:genetics / Q000378:metabolism*; D054648:Protein Phosphatase 2; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism*
D000952:Antigens, Polyomavirus Transforming; D016392:Proto-Oncogene Proteins pp60(c-src); D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2