Insulin receptor capping and its correlation with calmodulin-dependent myosin light chain kinase.
Both fluorescence microscopy and fluorometric analysis techniques have been used to characterize insulin receptor capping in IM-9 human lymphoblastoid cells. Morphologically, insulin caps appear similar to lectin or antiimmunoglobulin-induced caps displaying a preferential accumulation of actin, myosin, and actin-binding protein directly underneath the cap structure. Using the fluorescent calcium indicator quin2 we have detected no change in the calcium activity following insulin stimulation. However, in the presence of a number of calmodulin inhibitors, such as W-5, W-7, W-12, and trifluoperazine (TFP), insulin capping is significantly inhibited, which implies that a calmodulin-regulated process is involved. Using double immunofluorescence microscopy, we have found that the calmodulin-dependent myosin light chain kinase (MLCK) is concentrated directly beneath insulin caps. Upon treatment with trifluoperazine (TFP), the redistribution of both MLCK and insulin receptors are inhibited concomitantly. Our data indicate that the calmodulin-dependent myosin light chain kinase may be directly responsible for the activation of actomyosin-mediated contractility during insulin receptor capping.
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D000205:Actomyosin / Q000378:metabolism; D000634:Aminoquinolines; D002118:Calcium / Q000032:analysis; D002147:Calmodulin / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000648:ultrastructure*; D003208:Concanavalin A / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000032:analysis; D005470:Fluorometry; D006680:HLA Antigens / Q000276:immunology; D006801:Humans; D007074:Immunoglobulin G / Q000378:metabolism; D007328:Insulin / Q000378:metabolism; D037102:Lectins / Q000378:metabolism; D008214:Lymphocytes / Q000166:cytology; D008856:Microscopy, Fluorescence; D009219:Myosin-Light-Chain Kinase; D011494:Protein Kinases / Q000378:metabolism*; D011972:Receptor, Insulin / Q000378:metabolism*; D017452:Receptors, IgG; D011971:Receptors, Immunologic / Q000378:metabolism; D011975:Receptors, Mitogen / Q000378:metabolism; D013449:Sulfonamides / Q000494:pharmacology; D014018:Tissue Distribution; D014268:Trifluoperazine / Q000494:pharmacology; D014909:Wheat Germ Agglutinins
D000634:Aminoquinolines; D002147:Calmodulin; D003598:Cytoskeletal Proteins; D006680:HLA Antigens; D007074:Immunoglobulin G; D007328:Insulin; D037102:Lectins; D017452:Receptors, IgG; D011971:Receptors, Immunologic; D011975:Receptors, Mitogen; D013449:Sulfonamides; D014909:Wheat Germ Agglutinins; D003208:Concanavalin A; D014268:Trifluoperazine; C040184:W 12; C017967:W 7; C034700:N-(6-aminohexyl)-1-naphthalenesulfonamide; D000205:Actomyosin; D011494:Protein Kinases; D011972:Receptor, Insulin; D009219:Myosin-Light-Chain Kinase; C035739:Quin2; D002118:Calcium