Interaction of filamin A with the insulin receptor alters insulin-dependent activation of the mitogen-activated protein kinase pathway.
The biological actions of insulin are associated with a rapid reorganization of the actin cytoskeleton within cells in culture. Even though this event requires the participation of actin-binding proteins, the effect of filamin A (FLNa) on insulin-mediated signaling events is still unknown. We report here that human melanoma M2 cells lacking FLNa expression exhibited normal insulin receptor (IR) signaling, whereas FLNa-expressing A7 cells were unable to elicit insulin-dependent Shc tyrosine phosphorylation and p42/44 MAPK activation despite no significant defect in IR-stimulated phosphorylation of insulin receptor substrate-1 or activation of the phosphatidylinositol 3-kinase/AKT cascade. Insulin-dependent translocation of Shc, SOS1, and MAPK to lipid raft microdomains was markedly attenuated by FLNa expression. Coimmunoprecipitation experiments and in vitro binding assays demonstrated that FLNa binds constitutively to IR and that neither insulin nor depolymerization of actin by cytochalasin D affected this interaction. The colocalization of endogenous FLNa with IR was detected at the surface of HepG2 cells. Ectopic expression of a C-terminal fragment of FLNa (FLNaCT) in HepG2 cells blocked the endogenous IR-FLNa interaction and potentiated insulin-stimulated MAPK phosphorylation and transactivation of Elk-1 compared with vector-transfected cells. Expression of FLNaCT had no major effect on insulin-induced phosphorylation of the IR, insulin receptor substrate-1, or AKT, but it elicited changes in actin cytoskeletal structure and ruffle formation in HepG2 cells. Taken together, these results indicate that FLNa interacts constitutively with the IR to exert an inhibitory tone along the MAPK activation pathway.
Diabetes Section, Laboratory of Clinical Investigation, NIA, National Institutes of Health, Baltimore, Maryland 21224, USA.
Hua-Jun
HJ
He
Sutapa
S
Kole
Yong-Kook
YK
Kwon
Michael T
MT
Crow
Michel
M
Bernier
D001483:Base Sequence; D002460:Cell Line; D003285:Contractile Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004247:DNA / Q000235:genetics; D004268:DNA-Binding Proteins; D064448:Filamins; D006801:Humans; D066298:In Vitro Techniques; D007328:Insulin / Q000378:metabolism*; D055504:Insulin Receptor Substrate Proteins; D020935:MAP Kinase Signaling System / Q000502:physiology*; D021962:Membrane Microdomains / Q000378:metabolism; D008840:Microfilament Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D011972:Receptor, Insulin / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D014157:Transcription Factors; D015533:Transcriptional Activation; D014162:Transfection; D050807:ets-Domain Protein Elk-1
D003285:Contractile Proteins; D004268:DNA-Binding Proteins; C494112:ELK1 protein, human; D064448:Filamins; C526292:IRS1 protein, human; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D008840:Microfilament Proteins; D010446:Peptide Fragments; D010750:Phosphoproteins; D011518:Proto-Oncogene Proteins; D011994:Recombinant Proteins; D014157:Transcription Factors; D050807:ets-Domain Protein Elk-1; D004247:DNA; D011972:Receptor, Insulin