Enhancement of tyrosyl phosphorylation and protein expression of eps8 by v-Src

Two eps8 isoforms, p97(eps8) and p68(eps8), were previously identified as substrates for receptor tyrosine kinases. Analysis of eps8 phosphotyrosine content in v-Src transformed cells (IV5) revealed that both isoforms were highly tyrosyl phosphorylated and their readiness to be phosphorylated by Src in vitro further indicated that they were putative Src substrates as well. Indeed, the enhancement of tyrosyl phosphorylation of p97(eps8) detected in cells coexpressing both p97(eps8) and active Src relative to that in cells expressing p97(eps8) alone supported our hypothesis. The existence of common phosphotryptic peptides between in vitro ³²P-labeled p97(eps8) and p68(eps8) indicated that these two proteins shared the same Src-mediated sites. Further in vitro binding assays demonstrated that p68(eps8) was the major eps8 isoforms that could be precipitated by bacterial fusion protein containing Src SH3. Interestingly, both p68(eps8) and p97(eps8) were preferentially expressed in v-Src transformed cells and the presence of p68(eps8) appeared to depend on Src. Since p97(eps8) has been implicated in mitogenesis and tumorigenesis, its readiness to be phosphorylated and induced by v-Src might attribute to v-Src-mediated transformation. Copyright (C) 1999 Elsevier Science B.V.