A NOVEL 160-KDA PHOSPHOTYROSINE PROTEIN IN INSULIN-TREATED EMBRYONIC KIDNEY-CELLS IS A NEW MEMBER OF THE INSULIN-RECEPTOR SUBSTRATE FAMILY

We have previously identified a 160-kDa protein in human embryonic kid ney (HEK) 293 cells that undergoes rapid tyrosine phosphorylation in r esponse to insulin (PY160) (Kuhne, M. R., Zhao, Z., and Lienhard, G. E . (1995) Biochem. Biophys. Res. Commun. 211, 190-197). The phosphotyro sine form of PY160 was purified from insulin-treated HEK 293 cells by anti-phosphotyrosine immunoaffinity chromatography, the sequences of p eptides determined, and its cDNA cloned, The PY160 cDNA encodes a 1257 -amino acid protein that contains, in order from its N terminus, a ple ckstrin homology (PH) domain, a phosphotyrosine binding (PTB) domain, and, spread over the C-terminal portion, 12 potential tyrosine phospho rylation sites. Several of these sites are in motifs expected to bind specific SH2 domain-containing proteins: YXXM (7 sites), phosphatidyli nositol S-kinase; YVNM (1 site), Grb-2; and YIEV (1 site), either the protein-tyrosine phosphatase SHP-2 or phospholipase C gamma. Furthermo re, the PH and PTB domains are highly homologous (at least 40% identic al) to those found in insulin receptor substrates 1, 2, and 3 (IRS-1, IRS-2, and IRS-3). Thus, PY160 is a new member of the IRS family, whic h we have designated IRS-4.