The association between the SH2-containing inositol polyphosphate 5-Phosphatase 2 (SHIP2) and the adaptor protein APS has an impact on biochemical properties of both partners.
SHIP2 (SH2-containing inositol polyphosphate 5-phosphatase 2) is a phosphatidylinositol (3,4,5)-trisphosphate (PtdIns(3,4,5)P(3)) 5-phosphatase containing various motifs susceptible to mediate protein-protein interaction. In cell models, SHIP2 negatively regulates insulin signalling through its catalytic PtdIns(3,4,5)P(3) 5-phosphatase activity. We have previously reported that SHIP2 interacts with the c-Cbl associated protein (CAP) and c-Cbl, proteins implicated in the insulin cellular response regulating the small G protein TC10. The first steps of the TC10 pathway are the recruitment and tyrosine phosphorylation by the insulin receptor of the adaptor protein with Pleckstrin Homology and Src Homology 2 domains (APS). Herein, we show that SHIP2 can directly interact with APS in 3T3-L1 adipocytes and in transfected CHO-IR cells (Chinese hamster ovary cells stably transfected with the insulin receptor). Upon insulin stimulation, APS and SHIP2 are recruited to cell membranes as seen by immunofluorescence studies, which is consistent with their interaction. We also observed that SHIP2 negatively regulates APS insulin-induced tyrosine phosphorylation and consequently inhibits APS association with c-Cbl. APS, which specifically interacts with SHIP2, but not PTEN, in turn, increases the PtdIns(3,4,5)P(3) 5-phosphatase activity of SHIP2 in an inositol phosphatase assay. Co-transfection of SHIP2 and APS in CHO-IR cells further increases the inhibitory effect of SHIP2 on Akt insulin-induced phosphorylation. Therefore, the interaction between APS and SHIP2 provides to both proteins potential negative regulatory mechanisms to act on the insulin cascade.
Onnockx
Sheela
S
Institute of Interdisciplinary Research (IRIBHM), Brussels, Belgium.
De Schutter
Julie
J
Blockmans
Marianne
M
Xie
Jingwei
J
Jacobs
Christine
C
Vanderwinden
Jean-Marie
JM
Erneux
Christophe
C
Pirson
Isabelle
I
D041721:3T3-L1 Cells; D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D017667:Adipocytes / Q000378:metabolism; D019943:Amino Acid Substitution; D000818:Animals; D016466:CHO Cells; D019556:COS Cells; D020134:Catalytic Domain; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002522:Chlorocebus aethiops; D006224:Cricetinae; D003412:Cricetulus; D016650:Fluorescein-5-isothiocyanate; D019085:Fluorescent Antibody Technique, Direct; D005456:Fluorescent Dyes; D005982:Glutathione Transferase / Q000378:metabolism; D006639:Histidine / Q000737:chemistry; D006801:Humans; D000072181:Inositol Polyphosphate 5-Phosphatases; D007328:Insulin / Q000494:pharmacology; D051379:Mice; D032448:Myoblasts, Skeletal / Q000378:metabolism; D000072183:Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases; D010744:Phosphoric Monoester Hydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011233:Precipitin Tests; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011972:Receptor, Insulin / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012694:Serine / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D014162:Transfection; D014364:Tryptophan / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D005456:Fluorescent Dyes; D007328:Insulin; D011993:Recombinant Fusion Proteins; D012694:Serine; D006639:Histidine; D014364:Tryptophan; D005982:Glutathione Transferase; D011972:Receptor, Insulin; D010744:Phosphoric Monoester Hydrolases; D000072181:Inositol Polyphosphate 5-Phosphatases; C000606435:Inppl1 protein, mouse; D000072183:Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases; D016650:Fluorescein-5-isothiocyanate