The 145-kDa protein induced to associate with Shc by multiple cytokines is an inositol tetraphosphate and phosphatidylinositol 3,4,5-triphosphate 5-phosphatase.

Abstract

(3,4), and two SH2-containing adapter proteins, Grb2 and Shc, have been implicated in its activation. Specifically, these proteins have been shown to bind directly to tyrosine-phosphorylated receptors (5-7) or SH2 docking proteins (such as the insulin receptor substrate 1) (8). Grb2, a 25-kDa protein with two SH3 domains flanking one SH2 domain, shuttles the Ras guanine nucleotide exchange factor, Sosl, to activated receptors (or to insulin receptor substrate 1) (5, 7-11) so that Sosl can activate Ras by catalyzing the exchange of GDP for GTP (5, 9-11). Shc, another widely expressed protein that contains an N-terminal phosphotyrosine binding (PTB) domain (12-16) and a C-terminal SH2 domain (17), can associate, in its tyrosine-phosphorylated form, with Grb2-Sosl complexes and may increase Grb2-Sosl interactions after growth factor stimulation (9,18,19 MATERIALS AND METHODS Reagents. COS-cell-derived murine IL-3 and GM-CSF were produced as described (29). Rabbit antiserum to the Shcassociated p145 protein was generated by immunizing rabbits with a 15-mer obtained from amino acid sequencing (VPAE-GVSSLNEMINP) and crosslinked to keyhole limpet hemocyanin with glutaraldehyde (30). The glutathione S-transferase (GST) fusion proteins, consisting of the 27-kDa N-terminal and of GST linked to the Grb2 N-terminal (amino acid residues 5-56) or C-terminal (residues 163-215) SH3 domains of Grb2, were expressed in Escherichia coli in pGEX-2T plasmids (Pharmacia/LKB) and the fusion proteins were recovered from clarified lysates with glutathione (GSH)-agarose beads as described (31 Abbreviations: GM-CSF, granulocyte-macrophage colony-stimulating factor; GSH, glutathione; GST, glutathione S-transferase; IL-3, interleukin 3; NRS, normal rabbit serum; 5-ptase, inositol polyphosphate 5-phosphatase; Ptd1ns(3,4,5)P3, phosphatidylinositol 3,4,5-trisphosphate; Ins(1,3,4,5)P4, inositol 1,3,4,5-tetrakisphosphate; SH2, src homology 2 domain; PTB, phosphotyrosine binding; p145, the Shc-associated 145-kDa protein.

Cloning and characterization of APS, an adaptor molecule containing PH and SH2 domains that is tyrosine phosphorylated upon B-cell receptor stimulation

“…The APS-associated phosphoproteins were quite similar to those associated with Grb2 and c-Cbl. The major APS binding phosphoprotein, p145 was precipitated with all other antibodies, and it is probably SHIP which is reported to be associated with Shc and Grb2 (Damen et al, 1996;Osborne et al, 1996). A marginal amount of p120 c-Cbl was precipitated with APS.…”

Regulation of mast cells’ secretory response by co‐clustering the Type 1 Fcϵ receptor with the mast cell function‐associated antigen

“…These inhibitory receptors include amongst other the CTLA-4 and CD22 expressed by T and B cells, respectively, murine FccRIIB on B and mast cells or the killer cell inhibitory receptors (KIR) on natural killer (NK) cells [1,2,3]. These receptors, upon being co-clustered with the activating receptors undergo phosphorylation of their ITIM tyrosine residues and recruit SH2 domain containing phosphatases, which interfere with the activating receptors' stimulus-response coupling cascade [1,3,4,5]. The mast cells function-associated antigen (MAFA), another inhibitory receptor has a C-type lectin domain in its extracellular part and, like the other members of the above family, its intracellular domain contains an ITIM sequence [6].…”

Cloning and characterization of mPAL, a novel Shc SH2 domain-binding protein expressed in proliferating cells

“…A novel role for Shc has been suggested in which phosphorylation of Y239/Y240 leads to c-myc induction and suppression of apoptosis in Ba/F3 cells, in a Ras-independent manner (Gotoh et al, 1996). The Shc PTB domain binds directly to the cytoplasmic enzyme SHIP, an SH2 domain-containing inositol 5-phosphatase, in response to growth factor and cytokine stimulation in hematopoetic cells (Damen et al, 1996;Kavanaugh et al, 1996;Lioubin et al, 1996). Furthermore, proline-rich sequences in the Shc CH1 domain are proposed to mediate the interaction between Shc and the SH3 domain of eps8, a tyrosinephosphorylated protein involved in EGF receptormediated signaling (Matoskova et al, 1995;Bon®ni et al, 1996).…”