Coligation of the Fc receptor on B cells, Fc gamma RIIB1, with the B cell antigen receptor (BCR) leads to abortive BCR signaling. Here it was shown that the Fc gamma RIIB1 recruits the phosphotyrosine phosphatase PTP1C after BCR coligation. This association is mediated by the binding of a 13-amino acid tyrosine-phosphorylated sequence to the carboxyl-terminal Src homology 2 domain of PTP1C and activates PTP1C. Inhibitory signaling and PTP1C recruitment are dependent on the presence of the tyrosine within the 13-amino acid sequence. Inhibitory signaling mediated by Fc gamma RIIB1 is deficient in motheaten mice which do not express functional PTP1C. Thus, PTP1C is an effector of BCR-Fc gamma RIIB1 negative signal cooperativity.

Under resting conditions, the p85 regulatory subunit of phosphatidylinositol 3-kinase (PI3K) serves to both stabilize and inactivate the p110 catalytic subunit. The inhibitory activity of p85 is relieved by occupancy of the NH 2 -terminal SH2 domain of p85 by phosphorylated tyrosine. Src family kinases phosphorylate tyrosine 688 in p85, a process that we have shown to be reversed by the activity of the p85-associated SH2 domain-containing phosphatase SHP1. We demonstrate that phosphorylation of the downstream PI3K target Akt is increased in cells lacking SHP1, implicating phosphorylation of p85 in the regulation of PI3K activity. Furthermore, the in vitro specific activity of PI3K associated with tyrosinephosphorylated p85 is higher than that associated with nonphosphorylated p85. Expression of wild-type p85 inhibits PI3K enzyme activity as indicated by PI3K-dependent Akt phosphorylation. The inhibitory activity of p85 is accentuated by mutation of tyrosine 688 to alanine and reversed by mutation of tyrosine 688 to aspartic acid, changes that block and mimic tyrosine phosphorylation, respectively Strikingly, mutation of tyrosine 688 to aspartic acid completely reverses the inhibitory activity of p85 on cell viability and activation of the downstream targets Akt and NFB, indicative of the physiological relevance of p85 phosphorylation. Tyrosine phosphorylation of Tyr 688 or mutation of tyrosine 688 to aspartic acid is sufficient to allow binding to the NH 2 -terminal SH2 domain of p85. Thus an intramolecular interaction between phosphorylated Tyr 688 and the NH 2 -terminal SH2 domain of p85 can relieve the inhibitory activity of p85 on p110. Taken together, the data indicate that phosphorylation of Tyr 688 in p85 leads to a novel mechanism of PI3K regulation. The PI3K1 signaling cascade has been linked to proliferation, cell survival, differentiation, apoptosis, cytoskeletal rearrangement, and vacuolar trafficking. Growth factor-responsive Class IA PI3Ks consist of heterodimers of a 110-kDa catalytic subunit associated with an 85-kDa noncatalytic regulatory subunit designated p85. The p85 adapter subunits are encoded by at least three different genes with splice variation generating multiple proteins potentially serving many different functions (1). Of the known p85 adapter subunits and splice variants, nearly all contain two Src-homology 2 (SH2) domains, which enable p85 to bind phosphotyrosine in an appropriate amino acid context. The p85 SH2 domains most frequently, but not exclusively, recognize phosphotyrosine embedded in a YXXM motif (2). Most p85 gene products also include a Src homology 3 (SH3) domain, as well as other domains involved in protein-protein interactions (3). All p85 family members contain a p110-binding motif located between the two SH2 domains. The diversity of protein interaction domains found among p85 family members likely contributes to the ability of multiple signaling proteins and pathways to activate PI3K. Under resting conditions, p85 serves to both stabilize p110 protein and inhibi...

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