A highly phosphorylated subpopulation of insulin-like growth factor II/mannose 6-phosphate receptors is concentrated in a clathrin-enriched plasma membrane fraction.
Abstract
Insulin-like growth factor II (IGF-II)/mannose 6-phosphate (Man-6-P) receptors immunoprecipitated from purified plasma membranes of 32P-labeled rat adipocytes are markedly heterogenous in their phosphorylation state. Approximately 80% of the plasma membrane receptors are solubilized in 1% (vol/vol) Triton X-100 and are phosphorylated on serine residues at a stoichiometry of approximately 0.1-0.2 mol of phosphate per mol of receptor. In contrast, 15-20% of the receptors are Triton X-100-insoluble and are phosphorylated on serine and threonine residues at approximately 4 or 5 mol of phosphate per mol of receptor. This Triton X-100-insoluble membrane subfraction contains only 5% of the total plasma membrane protein and yet contains all of the clathrin heavy chain associated with plasma membrane, as detected by immunoblotting with a monoclonal antibody. Based on the relative yields of protein in the detergent-insoluble material, IGF-II/Man-6-P receptors are concentrated approximately equal to 3-fold in this clathrin-enriched subfraction. Insulin treatment of intact cells increased the total IGF-II/Man-6-P receptors in the Triton X-100-soluble fraction of the plasma membrane, whereas no change in receptor number in the detergent-insoluble fraction was seen. However, insulin markedly decreased the phosphorylation stoichiometry of the Triton X-100-insoluble receptors. Taken together, these results indicate that insulin decreases the phosphorylation state of a highly phosphorylated subpopulation of IGF-II/Man-6-P receptors on the plasma membrane. In addition, insulin action may prevent the concentration of these receptors in a clathrin-enriched membrane subfraction.
Full text
Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (1.3M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.
Images in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- MacDonald RG, Pfeffer SR, Coussens L, Tepper MA, Brocklebank CM, Mole JE, Anderson JK, Chen E, Czech MP, Ullrich A. A single receptor binds both insulin-like growth factor II and mannose-6-phosphate. Science. 1988 Mar 4;239(4844):1134–1137. [PubMed] [Google Scholar]
- Morgan DO, Edman JC, Standring DN, Fried VA, Smith MC, Roth RA, Rutter WJ. Insulin-like growth factor II receptor as a multifunctional binding protein. Nature. 1987 Sep 24;329(6137):301–307. [PubMed] [Google Scholar]
- von Figura K, Hasilik A. Lysosomal enzymes and their receptors. Annu Rev Biochem. 1986;55:167–193. [PubMed] [Google Scholar]
- Oka Y, Rozek LM, Czech MP. Direct demonstration of rapid insulin-like growth factor II Receptor internalization and recycling in rat adipocytes. Insulin stimulates 125I-insulin-like growth factor II degradation by modulating the IGF-II receptor recycling process. J Biol Chem. 1985 Aug 5;260(16):9435–9442. [PubMed] [Google Scholar]
- Mottola C, Czech MP. The type II insulin-like growth factor receptor does not mediate increased DNA synthesis in H-35 hepatoma cells. J Biol Chem. 1984 Oct 25;259(20):12705–12713. [PubMed] [Google Scholar]
- Hari J, Pierce SB, Morgan DO, Sara V, Smith MC, Roth RA. The receptor for insulin-like growth factor II mediates an insulin-like response. EMBO J. 1987 Nov;6(11):3367–3371. [PMC free article] [PubMed] [Google Scholar]
- Oka Y, Czech MP. Photoaffinity labeling of insulin-sensitive hexose transporters in intact rat adipocytes. Direct evidence that latent transporters become exposed to the extracellular space in response to insulin. J Biol Chem. 1984 Jul 10;259(13):8125–8133. [PubMed] [Google Scholar]
- Wardzala LJ, Simpson IA, Rechler MM, Cushman SW. Potential mechanism of the stimulatory action of insulin on insulin-like growth factor II binding to the isolated rat adipose cell. Apparent redistribution of receptors cycling between a large intracellular pool and the plasma membrane. J Biol Chem. 1984 Jul 10;259(13):8378–8383. [PubMed] [Google Scholar]
- Oppenheimer CL, Pessin JE, Massague J, Gitomer W, Czech MP. Insulin action rapidly modulates the apparent affinity of the insulin-like growth factor II receptor. J Biol Chem. 1983 Apr 25;258(8):4824–4830. [PubMed] [Google Scholar]
- Sahagian GG, Neufeld EF. Biosynthesis and turnover of the mannose 6-phosphate receptor in cultured Chinese hamster ovary cells. J Biol Chem. 1983 Jun 10;258(11):7121–7128. [PubMed] [Google Scholar]
- Corvera S, Czech MP. Mechanism of insulin action on membrane protein recycling: a selective decrease in the phosphorylation state of insulin-like growth factor II receptors in the cell surface membrane. Proc Natl Acad Sci U S A. 1985 Nov;82(21):7314–7318. [PMC free article] [PubMed] [Google Scholar]
- Willingham MC, Pastan IH, Sahagian GG, Jourdian GW, Neufeld EF. Morphologic study of the internalization of a lysosomal enzyme by the mannose 6-phosphate receptor in cultured Chinese hamster ovary cells. Proc Natl Acad Sci U S A. 1981 Nov;78(11):6967–6971. [PMC free article] [PubMed] [Google Scholar]
- Pearse BM. Coated vesicles from human placenta carry ferritin, transferrin, and immunoglobulin G. Proc Natl Acad Sci U S A. 1982 Jan;79(2):451–455. [PMC free article] [PubMed] [Google Scholar]
- RODBELL M. METABOLISM OF ISOLATED FAT CELLS. I. EFFECTS OF HORMONES ON GLUCOSE METABOLISM AND LIPOLYSIS. J Biol Chem. 1964 Feb;239:375–380. [PubMed] [Google Scholar]
- Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. [PubMed] [Google Scholar]
- Oppenheimer CL, Pessin JE, Massague J, Gitomer W, Czech MP. Insulin action rapidly modulates the apparent affinity of the insulin-like growth factor II receptor. J Biol Chem. 1983 Apr 25;258(8):4824–4830. [PubMed] [Google Scholar]
- Towbin H, Staehelin T, Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. [PMC free article] [PubMed] [Google Scholar]
- Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. [PubMed] [Google Scholar]
- Bruder G, Wiedenmann B. Identification of a distinct 9S form of soluble clathrin in cultured cells and tissues. Exp Cell Res. 1986 Jun;164(2):449–462. [PubMed] [Google Scholar]
- Brown WJ, Farquhar MG. The mannose-6-phosphate receptor for lysosomal enzymes is concentrated in cis Golgi cisternae. Cell. 1984 Feb;36(2):295–307. [PubMed] [Google Scholar]
- Geuze HJ, Slot JW, Strous GJ, Hasilik A, Von Figura K. Ultrastructural localization of the mannose 6-phosphate receptor in rat liver. J Cell Biol. 1984 Jun;98(6):2047–2054. [PMC free article] [PubMed] [Google Scholar]
- Pearse BM. Assembly of the mannose-6-phosphate receptor into reconstituted clathrin coats. EMBO J. 1985 Oct;4(10):2457–2460. [PMC free article] [PubMed] [Google Scholar]
- Keen JH, Chestnut MH, Beck KA. The clathrin coat assembly polypeptide complex. Autophosphorylation and assembly activities. J Biol Chem. 1987 Mar 15;262(8):3864–3871. [PubMed] [Google Scholar]
- Usami M, Takahashi A, Kadota K. Protein kinase and its endogenous substrates in coated vesicles. Biochim Biophys Acta. 1984 Apr 24;798(3):306–312. [PubMed] [Google Scholar]
- Bar-Zvi D, Branton D. Clathrin-coated vesicles contain two protein kinase activities. Phosphorylation of clathrin beta-light chain by casein kinase II. J Biol Chem. 1986 Jul 25;261(21):9614–9621. [PubMed] [Google Scholar]
- Corvera S, Roach PJ, DePaoli-Roach AA, Czech MP. Insulin action inhibits insulin-like growth factor-II (IGF-II) receptor phosphorylation in H-35 hepatoma cells. IGF-II receptors isolated from insulin-treated cells exhibit enhanced in vitro phosphorylation by casein kinase II. J Biol Chem. 1988 Mar 5;263(7):3116–3122. [PubMed] [Google Scholar]












