THE SRC HOMOLOGY-2 (SH2) DOMAIN OF SH2-CONTAINING INOSITOL PHOSPHATASE (SHIP) IS ESSENTIAL FOR TYROSINE PHOSPHORYLATION OF SHIP, ITS ASSOCIATION WITH SHC, AND ITS INDUCTION OF APOPTOSIS

In this study we have investigated the role that the Src homology 2 do main (SH2) of the 145-kDa 5-phosphatase, SH2-containing inositol phosp hatase (SHIP), plays in three of the properties that have been associa ted with this protein following cytokine stimulation: its association with Shc, its tyrosine phosphorylation, and its inhibition of hemopoie tic cell growth. In vitro studies using this SH2 domain revealed that it was capable of binding directly to the Tyr(P)(317) motif of Shc wit h a K-D of approximately 290 nM, in keeping with other specific SH2/Ty r(P) interactions. In vivo analysis revealed the SH2 and NPXpY motifs of SHIP acted together, with the Tyr(P)(317),and phosphotyrosine bindi ng (PTB) domains of Shc, respectively, to ensure a high affinity SHIP Shc complex. Expression of cDNAs encoding hemagglutinin-tagged wild ty pe and SH2-inactivated forms of SHIP in the murine hemopoietic cell li ne DA-ER revealed that wild type SHIP becomes both tyrosine-phosphoryl ated and associated with Shc following interleukin-3 stimulation, as e xpected, but the SH2-inactivated SHIPs do neither. Moreover, while the growth rates of parental DA-ER cells and cells expressing these vario us SHIP constructs are identical, the wild type SHIP-expressing cells die, via programmed cell death, far more rapidly than parental cells. Cells expressing SH2-inactivated SHIPs, on the other hand, show either a reduced or no effect on apoptosis, These results suggest that the S H2 domain of SHIP is required not only for the tyrosine phosphorylatio n of SHIP and Shc association following cytokine stimulation but also for its induction of apoptosis.