Association between src-kinases and the polyoma virus oncogene middle T-antigen requires PP2A and a specific sequence motif

Polyoma virus encodes a potent oncogene, the middle T-antigen (MT), that induces cell transformation by copying the actions of tyrosine kinase associated growth factor receptors, A crucial component of MT transformation is its ability to bind and stimulate the activity of src-family kinases. However, the mechanism by which this is achieved remains unclear. Tyrosine phosphorylation of MT by sr c-kinases then provides binding sites for SH2 and PTB domain containing molecules in a paradigm of receptor action. We present evidence here that the MT/src complex contains equi-molar amounts of PP2A, and that phosphatase activity may be required for the interaction of MT with both PP2A and the src-family. PP2A, then, is a necessary component of the;MT-src complex. We also show that tno motifs in the 185 to 210 region of MT, each consisting of a basic area followed by a serine or threonine, are essential for interaction with sr c-kinases, but not PP2A. The spacing between the serine or threonine and the basic sequence also appears to be important. Substituting a cysteine residue in place of Thr203 in MT has no affect on the binding of pp60(c-src), showing that these sites interact with src-kinases by a novel mechanism that does not require phosphorylation.