Phosphorylation of the adaptor protein SH2B1? regulates its ability to enhance growth hormone-dependent macrophage motility.
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ABSTRACT: Previous studies have shown that growth hormone (GH) recruits the adapter protein SH2B1? to the GH-activated, GH receptor-associated tyrosine kinase JAK2, implicating SH2B1? in GH-dependent actin cytoskeleton remodeling, and suggesting that phosphorylation at serines 161 and 165 in SH2B1? releases SH2B1? from the plasma membrane. Here, we examined the role of SH2B1? in GH regulation of macrophage migration. We show that GH stimulates migration of cultured RAW264.7 macrophages, and primary cultures of peritoneal and bone marrow-derived macrophages. SH2B1? overexpression enhances, whereas SH2B1 knockdown inhibits, GH-dependent motility of RAW macrophages. At least two independent mechanisms regulate the SH2B1?-mediated changes in motility. In response to GH, tyrosines 439 and 494 in SH2B1? are phosphorylated. Mutating these tyrosines in SH2B1? decreases both basal and GH-stimulated macrophage migration. In addition, mutating the polybasic nuclear localization sequence (NLS) in SH2B1? or creating the phosphomimetics SH2B1?(S161E) or SH2B1?(S165E), all of which release SH2B1? from the plasma membrane, enhances macrophage motility. Conversely, SH2B1?(S161/165A) exhibits increased localization at the plasma membrane and decreased macrophage migration. Mutating the NLS or the nearby serine residues does not alter GH-dependent phosphorylation on tyrosines 439 and 494 in SH2B1?. Mutating tyrosines 439 and 494 does not affect localization of SH2B1? at the plasma membrane or movement of SH2B1? into focal adhesions. Taken together, these results suggest that SH2B1? enhances GH-stimulated macrophage motility via mechanisms involving phosphorylation of SH2B1? on tyrosines 439 and 494 and movement of SH2B1? out of the plasma membrane (e.g. as a result of phosphorylation of serines 161 and 165).
SUBMITTER: Su HW
PROVIDER: S-EPMC3678408 | biostudies-literature | 2013 Apr
REPOSITORIES: biostudies-literature
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