Ca2+ regulates calmodulin binding to IQ motifs in IRS-1.
IRS-proteins couple the receptors for insulin and various cytokines to signalling proteins containing Src homology 2 (SH2) domains. Here we demonstrate that calmodulin, a mediator of Ca(2+)-dependent physiological processes, associates with IRS-1 in a phosphotyrosine-independent manner. IRS-1 coimmunoprecipitated with calmodulin from lysates of Chinese hamster ovary cells expressing IRS-1. The interaction was modulated by Ca2+, and calmodulin binding to IRS-1 was enhanced by increasing intracellular Ca2+ with A23187. In contrast, trifluoperazine, a cell-permeable calmodulin antagonist, decreased binding of calmodulin to IRS-1. Insulin stimulated tyrosine phosphorylation of IRS-1, but did not significantly alter the interaction between calmodulin and IRS-1. IQ-like motifs occur between residues 106-126 and 839-859 of IRS-1. Synthetic peptides based on the these sequences inhibited the association between IRS-1 and calmodulin. These data demonstrate that calmodulin binds to IRS-1 in intact cells in a Ca(2+)-regulated manner, providing a molecular link between the signalling pathways.
Munshi
H G
HG
Department of Pathology, Brigham and Women's Hospital, Boston, Massachusetts, USA.
Burks
D J
DJ
Joyal
J L
JL
White
M F
MF
Sacks
D B
DB
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000378:metabolism; D001665:Binding Sites; D016466:CHO Cells; D002118:Calcium / Q000494:pharmacology*; D002147:Calmodulin / Q000276:immunology / Q000378:metabolism*; D016384:Consensus Sequence / Q000235:genetics; D006224:Cricetinae; D004591:Electrophoresis, Polyacrylamide Gel; D007328:Insulin / Q000494:pharmacology; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism; D010750:Phosphoproteins / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D010766:Phosphorylation; D011233:Precipitin Tests; D011485:Protein Binding / Q000187:drug effects; D051381:Rats; D014268:Trifluoperazine / Q000494:pharmacology; D014443:Tyrosine / Q000378:metabolism
D000911:Antibodies, Monoclonal; D002147:Calmodulin; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins; C526295:Irs1 protein, mouse; C526296:Irs1 protein, rat; C526298:Irs2 protein, mouse; C526299:Irs2 protein, rat; D010446:Peptide Fragments; D010750:Phosphoproteins; D014268:Trifluoperazine; D014443:Tyrosine; D002118:Calcium