Tyrosine phosphorylation of tub and its association with Src homology 2 domain-containing proteins implicate tub in intracellular signaling by insulin.
A mutation in the tub gene leads to maturity-onset obesity, insulin resistance, and progressive retinal and cochlear degeneration in mice. tub is a member of a growing family of genes that encode proteins of unknown function that are remarkably conserved across species. The absence of obvious transmembrane domain(s) or signal sequence peptide motif(s) suggests that Tub is an intracellular protein. Additional sequence analysis revealed the presence of putative tyrosine phosphorylation motifs and Src homology 2 (SH2)-binding sites. Here we demonstrate that in CHO-IR cells, transfected Tub is phosphorylated on tyrosine in response to insulin and insulin-like growth factor-1 and that in PC12 cells, insulin but not EGF induced tyrosine phosphorylation of endogenous Tub. In vitro, Tub is phosphorylated by purified insulin receptor kinase as well as by Abl and JAK 2 but not by epidermal growth factor receptor and Src kinases. Furthermore, upon tyrosine phosphorylation, Tub associated selectively with the SH2 domains of Abl, Lck, and the C-terminal SH2 domain of phospholipase Cgamma and insulin enhanced the association of Tub with endogenous phospholipase Cgamma in CHO-IR cells. These data suggest that Tub may function as an adaptor protein linking the insulin receptor, and possibly other protein-tyrosine kinases, to SH2-containing proteins.
Kapeller
R
R
Millennium Pharmaceuticals, Inc., Cambridge, Massachusetts 02139, USA. kapeller@mpi.com
Moriarty
A
A
Strauss
A
A
Stubdal
H
H
Theriault
K
K
Siebert
E
E
Chickering
T
T
Morgenstern
J P
JP
Tartaglia
L A
LA
Lillie
J
J
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D007328:Insulin / Q000378:metabolism*; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D053614:Janus Kinase 2; D016374:Oncogene Proteins v-abl / Q000378:metabolism; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism*; D011485:Protein Binding; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism; D011972:Receptor, Insulin / Q000378:metabolism; D015398:Signal Transduction; D014162:Transfection; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D007328:Insulin; D016374:Oncogene Proteins v-abl; D011506:Proteins; D011518:Proto-Oncogene Proteins; C098988:Tub protein, mouse; D019000:Phosphotyrosine; D007334:Insulin-Like Growth Factor I; D011505:Protein-Tyrosine Kinases; D020794:Receptor Protein-Tyrosine Kinases; D011972:Receptor, Insulin; C507926:Jak2 protein, mouse; D053614:Janus Kinase 2