Phosphatase 2A associated with polyomavirus small‐T or middle‐T antigen is an okadaic acid‐sensitive tyrosyl phosphatase

Abstract

Papovavirus tumor antigens have been shown to associate with the cellular phosphoserinehreonine-specific protein phosphatase 2A (PP2A). We were interested in the consequences that Tantigen association might have on PP2A activity and so studies of the phosphatase activity in immunoprecipitates, prepared from polyoma virus-transformed or polyoma virus-infected mouse 3T3 fibroblasts, were performed. The phosphoserine/threonine phosphatase activity, measured with phosphorylase a as the substrate, showed all the characteristics of PP2A. It was stimulated by polycations, inhibited by fluoride or p-nitrophenyl phosphate, sensitive to okadaic acid and microcystin and insensitive to inhibitor-1 and inhibitor-2. Phosphotyrosyl phosphatase (PTPase) activity was associated with the middle-T/small-T-associated complex when reduced, carboxamidomethylated and maleylated lysozyme, phosphorylated exclusively on tyrosyl residues, was used as the substrate. This PTPase activity was as sensitive to okadaic acid as was the phosphorylase phosphatase activity ; it could be inhibited by phosphorylase a and did not dephosphorylate poly(G1u8nTyr2n). The level of middle-T/small-T-associated PTPase activity relative to the phosphorylase phosphatase activity was tenfold higher than that of the purified dimeric PP2A. A similar activity ratio was observed with the purified phosphatase after stimulation with a cellular protein, designated phosphotyrosyl phosphatase activator. These results suggest that the same enzyme may possess dual specificity. In contrast to the cellular trimeric PP2A, containing the 55-kDa putative regulatory subunit, the middle-Thmall-T-associated enzyme had low activity towards a retinoblastoma peptide phosphorylated by p3Pdc2. These results indicate how middle-Thmall-T might effect the activity of PP2A in polyoma virus-transformed cells.

Protein phosphatase 2A subunit assembly: the catalytic subunit carboxy terminus is important for binding cellular B subunit but not polyomavirus middle tumor antigen

“…Cayla and coworkers had previously analysed immunoprecipitates of an undetermined mixture of polyomavirus middle and small tumor antigens (Cayla et al, 1993). They found that the ratio of PP2A activity in these immunoprecipitates towards phosphorylase a and a cdc2-phosphorylated retinoblastoma protein peptide was similar to that obtained for biochemically puri®ed A/C heterodimer.…”

Biochemical characterization of recombinant yeast PPZ1, a protein phosphatase involved in salt tolerance

“…The preparation and phosphorylation of the substrate and the assay conditions were as described [23].…”

Methylation of the Protein Phosphatase 2A Catalytic Subunit Is Essential for Association of Bα Regulatory Subunit But Not SG2NA, Striatin, or Polyomavirus Middle Tumor Antigen

“…Moreover, a family of papovavirus PP2A regulatory subunits exists that includes simian virus 40 and polyomavirus small tumor antigens and polyomavirus middle tumor antigen (MT) (Pallas et al, 1990). In all these cases, the PP2A core A/C heterodimer is found complexed with the regulatory subunit, and the substrate specificity and activity of PP2A are modulated (Yang et al, 1991;Cayla et al, 1993;Cegielska et al, 1994;Kamibayashi et al, 1994;Mayer-Jaekel et al, 1994;Ogris et al, 1997;Moreno et al, 2000).…”