PHOSPHORYLATION OF ETS1 REGULATES THE COMPLEMENTATION OF A CSF-1 RECEPTOR IMPAIRED IN MITOGENESIS

Ets1, the founder member of the Ets transcription factor family, is in volved in a variety of developmental and cellular processes, Previous studies have shown that serine phosphorylation of Ets1 inhibits its DN A binding activity, suggesting that phosphorylation is important in th e regulation of Ets1 function. To further examine Ets1 phosphorylation , we ectopically expressed Ets1 in fibroblasts and stimulated these ce lls with serum, Using two-dimensional tryptic phosphopeptide analysis and site-directed mutagenesis, we found that Ets1 was phosphorylated o n threonine 38, a residue conserved in several Ets proteins, Substitut ion of this residue with alanine enhanced CSF-1-dependent colony forma tion in semi-solid medium of NIH3T3 cells expressing a mitogenically d efective CSF-1 receptor [Y809F], Threonine 38 is part of a consensus a mino-acid sequence frequently recognized and targeted by members of th e MAP kinase family, Moreover, this residue is phosphorylated in vitro by recombinant ERK2, which suggests that the kinase which phosphoryla tes threonine 38 in vivo is a member of the MAP kinase family, In addi tion, phosphorylation on threonine 38 seems to negatively regulate Ets 1 activity in response to growth-factor stimulation.