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Regulation of SH2 domain‐containing inositol 5′ phosphatase by the cyclic AMP dependent protein kinase
Author(s) -
Zhang Jun,
Garrison James C.
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.690.2
Subject(s) - phosphorylation , phosphatase , inositol , phosphatidylinositol , protein kinase a , chemistry , kinase , gq alpha subunit , biochemistry , microbiology and biotechnology , signal transduction , receptor , biology , g protein
The SH2 domain‐containing inositol 5' phosphatase, SHIP1, regulates hematopoietic cell function by opposing the action of phosphatidylinositol 3‐kinase (PI3K) and maintains low levels of phosphatidylinositol‐3,4,5‐trisphosphate (PIP 3 ). The activation of the cyclic AMP dependent protein kinase (PKA) opposes many of the pro‐inflammatory responses of hematopoietic cells. Thus, we tested to see if the activity of SHIP1 was regulated via phosphorylation by PKA. We prepared highly pure SHIP1 from HEK‐293 cells by immunoaffinity chromatography. Purified SHIP1 can be rapidly phosphorylated by PKA to a stoichiometry of over 0.6 mole PO 4 /mole SHIP1. In 32 P labeled HEK‐293 cells transfected with SHIP1, stimulation with Sp‐cAMPS or activation of the endogenous β‐adrenergic receptor caused an increase in the phosphorylation state of SHIP1. Inhibition of protein phosphatase activity with okadaic acid also increased the phosphorylation of SHIP1. The 5' phosphatase activity of SHIP1 was measured directly using an improved Malachite Green assay. Phosphorylation of the protein in vitro or in cells increased the activity of SHIP1 two fold. Thus, activation of G‐protein coupled receptors which raise cyclic AMP cause SHIP1 to be phosphorylated and stimulate its inositol phosphatase activity. These results outline a novel mechanism of SHIP1 activation by PKA phosphorylation. Supported by NIH Grant R01GM076236.