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. 1981 Oct 1;91(1):17–25. doi: 10.1083/jcb.91.1.17

Surface redistribution of 125I-insulin in cultured human lymphocytes

PMCID: PMC2111954  PMID: 7028760

Abstract

The cultured human lymphocyte (IM-9) binds 125I-insulin by a receptor- mediated process; the receptor, in turn, is regulated by the ligand. In the present study we have examined quantitatively the morphologic events involved in 125I-insulin interaction with the surface of the lymphocyte. At 2 min of incubation of 15 degrees or 37 degrees C, the ligand localizes preferentially at the villous surface of the cell, whereas with longer periods of incubation, the ligand distributes indistinguishably between the villous and nonvillous surface. When rebinding is blocked, 125I-insulin localizes preferentially at the nonvillous surface of the cell. When the total cell surface is considered, there is little preferential association with coated pits; when only the nonvillous surface is considered, a preferential association with coated pits is found and is quantitatively increased in the absence of rebinding of the ligand. This cell has an abundant villous surface (approximately 55% of the total surface); and, as seen on freeze-fracture replicas, the plasma membrane of the villous surface contains a 60% greater density of intramembrane particles than the nonvillous surface. The data suggest an ordered pattern of insulin interaction with the cell surface (i.e., binding to villi followed by redistribution to the nonvillous portion of the cell containing coated pits). These events probably reflect the mechanism by which the cell segregates specific receptors and related proteins in the plane of the membrane so that they can be selectively removed.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anderson R. G., Brown M. S., Goldstein J. L. Role of the coated endocytic vesicle in the uptake of receptor-bound low density lipoprotein in human fibroblasts. Cell. 1977 Mar;10(3):351–364. doi: 10.1016/0092-8674(77)90022-8. [DOI] [PubMed] [Google Scholar]
  2. Barazzone P., Carpentier J. L., Gorden P., Van Obberghen E., Orci L. Polar redistribution of 125I-labelled insulin on the plasma membrane of cultured human lymphocytes. Nature. 1980 Jul 24;286(5771):401–403. doi: 10.1038/286401a0. [DOI] [PubMed] [Google Scholar]
  3. Bergeron J. J., Levine G., Sikstrom R., O'Shaughnessy D., Kopriwa B., Nadler N. J., Posner B. I. Polypeptide hormone binding sites in vivo: initial localization of 125I-labeled insulin to hepatocyte plasmalemma as visualized by electron microscope radioautography. Proc Natl Acad Sci U S A. 1977 Nov;74(11):5051–5055. doi: 10.1073/pnas.74.11.5051. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bilheimer D. W., Ho Y. K., Brown M. S., Anderson R. G., Goldstein J. L. Genetics of the low density lipoprotein receptor. Diminished receptor activity in lymphocytes from heterozygotes with familial hypercholesterolemia. J Clin Invest. 1978 Mar;61(3):678–696. doi: 10.1172/JCI108980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Carpentier J. L., Gorden P., Amherdt M., Van Obberghen E., Kahn C. R., Orci L. 125I-insulin binding to cultured human lymphocytes. Initial localization and fate of hormone determined by quantitative electron microscopic autoradiography. J Clin Invest. 1978 Apr;61(4):1057–1070. doi: 10.1172/JCI109005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Carpentier J. L., Gorden P., Freychet P., Le Cam A., Orci L. Lysosomal association of internalized 125I-insulin in isolated rat hepatocytes. Direct demonstration by quantitative electron microscopic autoradiography. J Clin Invest. 1979 Jun;63(6):1249–1261. doi: 10.1172/JCI109420. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Carpentier J. L., Gorden P., Goldstein J. L., Anderson R. G., Brown M. S., Orci L. Binding and internalization of 125I-LDL in normal and mutant human fibroblasts. A quantitative autoradiographic study. Exp Cell Res. 1979 Jun;121(1):135–142. doi: 10.1016/0014-4827(79)90453-1. [DOI] [PubMed] [Google Scholar]
  8. Carpentier J. L., Perrelet A., Orci L. Perturbation of the adipose cell plasma membrane in obese (ob/ob) mice. Lab Invest. 1977 Sep;37(3):260–264. [PubMed] [Google Scholar]
  9. Carpentier J. L., van Obberghen E., Gordon P., Orci L. Binding, membrane redistribution, internalization and lysosomal association of [125I]anti-insulin receptor antibody in IM-9-cultured human lymphocyte. A comparison with [125I]insulin. Exp Cell Res. 1981 Jul;134(1):81–92. doi: 10.1016/0014-4827(81)90465-1. [DOI] [PubMed] [Google Scholar]
  10. Carpentier J., Perrelet A., Orci L. Morphological changes of the adipose cell plasma membrane during lipolysis. J Cell Biol. 1977 Jan;72(1):104–117. doi: 10.1083/jcb.72.1.104. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Flier J. S., Kahn C. R., Roth J., Bar R. S. Antibodies that impair insulin receptor binding in an unusual diabetic syndrome with severe insulin resistance. Science. 1975 Oct 3;190(4209):63–65. doi: 10.1126/science.170678. [DOI] [PubMed] [Google Scholar]
  12. Ghose K., Turner P., Grant I. N. The assessment of interactions in man between tyramine and viloxazine hydrochloride, an antidepressant drug. Br J Clin Pharmacol. 1976 Aug;3(4):668–671. doi: 10.1111/j.1365-2125.1976.tb04893.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Goldfine I. D., Jones A. L., Hradek G. T., Wong K. Y., Mooney J. S. Entry of insulin into human cultured lymphocytes: electron microscope autoradiographic analysis. Science. 1978 Nov 17;202(4369):760–763. doi: 10.1126/science.715440. [DOI] [PubMed] [Google Scholar]
  14. Gorden P., Carpentier J. L., Cohen S., Orci L. Epidermal growth factor: morphological demonstration of binding, internalization, and lysosomal association in human fibroblasts. Proc Natl Acad Sci U S A. 1978 Oct;75(10):5025–5029. doi: 10.1073/pnas.75.10.5025. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gorden P., Carpentier J. L., Freychet P., LeCam A., Orci L. Intracellular translocation of iodine-125-labeled insulin: direct demonstration in isolated hepatocytes. Science. 1978 May 19;200(4343):782–785. doi: 10.1126/science.644321. [DOI] [PubMed] [Google Scholar]
  16. Gordon P., Carpentier J. L., Van Obberghen E., Barazzone P., Roth J., Orci L. Insulin-induced receptor loss in the cultured human lymphocyte: quantitative morphological perturbations in the cell and plasma membrane. J Cell Sci. 1979 Oct;39:77–88. doi: 10.1242/jcs.39.1.77. [DOI] [PubMed] [Google Scholar]
  17. Grant C. W., McConnell H. M. Glycophorin in lipid bilayers. Proc Natl Acad Sci U S A. 1974 Dec;71(12):4653–4657. doi: 10.1073/pnas.71.12.4653. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Grinnell F., Tobleman M. Q., Hackenbrock C. R. The distribution and mobility of anionic sites on the surfaces of baby hamster kidney cells. J Cell Biol. 1975 Sep;66(3):470–479. doi: 10.1083/jcb.66.3.470. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. HUNTER W. M., GREENWOOD F. C. Preparation of iodine-131 labelled human growth hormone of high specific activity. Nature. 1962 May 5;194:495–496. doi: 10.1038/194495a0. [DOI] [PubMed] [Google Scholar]
  20. Haigler H. T., McKanna J. A., Cohen S. Direct visualization of the binding and internalization of a ferritin conjugate of epidermal growth factor in human carcinoma cells A-431. J Cell Biol. 1979 May;81(2):382–395. doi: 10.1083/jcb.81.2.382. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hammons G. T., Jarett L. Lysosomal degradation of receptor-bound 125I-labeled insulin by rat adipocytes: its characterization and dissociation from the short-term biologic effects of insulin. Diabetes. 1980 Jun;29(6):475–486. doi: 10.2337/diab.29.6.475. [DOI] [PubMed] [Google Scholar]
  22. Helenius A., Kartenbeck J., Simons K., Fries E. On the entry of Semliki forest virus into BHK-21 cells. J Cell Biol. 1980 Feb;84(2):404–420. doi: 10.1083/jcb.84.2.404. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Jarett L., Smith R. M. Ultrastructural localization of insulin receptors on adipocytes. Proc Natl Acad Sci U S A. 1975 Sep;72(9):3526–3530. doi: 10.1073/pnas.72.9.3526. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Jarrett D. B., Roth J., Kahn C. R., Flier J. S. Direct method for detection and characterization of cell surface receptors for insulin by means of 125I-labeled autoantibodies against the insulin receptor. Proc Natl Acad Sci U S A. 1976 Nov;73(11):4115–4119. doi: 10.1073/pnas.73.11.4115. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kahn C. R., Baird K., Filier J. S., Jarrett D. B. Effects of autoantibodies to the insulin receptor on isolated adipocytes. Studies of insulin binding and insulin action. J Clin Invest. 1977 Nov;60(5):1094–1106. doi: 10.1172/JCI108861. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Lindstrom J., Anholt R., Einarson B., Engel A., Osame M., Montal M. Purification of acetylcholine receptors, reconstitution into lipid vesicles, and study of agonist-induced cation channel regulation. J Biol Chem. 1980 Sep 10;255(17):8340–8350. [PubMed] [Google Scholar]
  27. Luborsky J. L., Behrman H. R. Localization of LH receptors on luteal cells with a ferritin--LH conjugate. Mol Cell Endocrinol. 1979 Aug;15(2):61–78. doi: 10.1016/0303-7207(79)90008-x. [DOI] [PubMed] [Google Scholar]
  28. Maxfield F. R., Schlessinger J., Shechter Y., Pastan I., Willingham M. C. Collection of insulin, EGF and alpha2-macroglobulin in the same patches on the surface of cultured fibroblasts and common internalization. Cell. 1978 Aug;14(4):805–810. doi: 10.1016/0092-8674(78)90336-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Orci L., Carpentier J. L., Perrelet A., Anderson R. G., Goldstein J. L., Brown M. S. Occurrence of low density lipoprotein receptors within large pits on the surface of human fibroblasts as demonstrated by freeze-etching. Exp Cell Res. 1978 Apr;113(1):1–13. doi: 10.1016/0014-4827(78)90081-2. [DOI] [PubMed] [Google Scholar]
  30. Orci L., Rufener C., Malaisse-Lagae F., Blondel B., Amherdt M., Bataille D., Freychet P., Perrelet A. Symposium III: Hormones and hormone receptors. A morphological approach to surface receptors in islet and liver cells. Isr J Med Sci. 1975 Jul;11(7):639–655. [PubMed] [Google Scholar]
  31. Salpeter M. M., McHenry F. A., Salpeter E. E. Resolution of electron microscope autoradiography. IV. Application to analysis of autoradiographs. J Cell Biol. 1978 Jan;76(1):127–145. doi: 10.1083/jcb.76.1.127. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Schlessinger J., Shechter Y., Willingham M. C., Pastan I. Direct visualization of binding, aggregation, and internalization of insulin and epidermal growth factor on living fibroblastic cells. Proc Natl Acad Sci U S A. 1978 Jun;75(6):2659–2663. doi: 10.1073/pnas.75.6.2659. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Schlessinger J., Van Obberghen E., Kahn C. R. Insulin and antibodies against insulin receptor cap on the membrane of cultured human lymphocytes. Nature. 1980 Aug 14;286(5774):729–731. doi: 10.1038/286729a0. [DOI] [PubMed] [Google Scholar]
  34. Silva P. P., Nicolson G. L. Freeze-etch localization of concanavalin A receptors to the membrane intercalated particles of human erythrocyte ghost membranes. Biochim Biophys Acta. 1974 Sep 23;363(3):311–319. doi: 10.1016/0005-2736(74)90071-6. [DOI] [PubMed] [Google Scholar]
  35. Tillack T. W., Scott R. E., Marchesi V. T. The structure of erythrocyte membranes studied by freeze-etching. II. Localization of receptors for phytohemagglutinin and influenza virus to the intramembranous particles. J Exp Med. 1972 Jun 1;135(6):1209–1227. doi: 10.1084/jem.135.6.1209. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Weibel E. R. Stereological principles for morphometry in electron microscopic cytology. Int Rev Cytol. 1969;26:235–302. doi: 10.1016/s0074-7696(08)61637-x. [DOI] [PubMed] [Google Scholar]
  37. Willingham M. C., Maxfield F. R., Pastan I. H. alpha 2 Macroglobulin binding to the plasma membrane of cultured fibroblasts. Diffuse binding followed by clustering in coated regions. J Cell Biol. 1979 Sep;82(3):614–625. doi: 10.1083/jcb.82.3.614. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Yu J., Branton D. Reconstitution of intramembrane particles in recombinants of erythrocyte protein band 3 and lipid: effects of spectrin-actin association. Proc Natl Acad Sci U S A. 1976 Nov;73(11):3891–3895. doi: 10.1073/pnas.73.11.3891. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. de Petris S. Preferential distribution of surface immunoglobulins on microvilli. Nature. 1978 Mar 2;272(5648):66–68. doi: 10.1038/272066a0. [DOI] [PubMed] [Google Scholar]
  40. de Petris S., Raff M. C. Normal distribution, patching and capping of lymphocyte surface immunoglobulin studied by electron microscopy. Nat New Biol. 1973 Feb 28;241(113):257–259. doi: 10.1038/newbio241257a0. [DOI] [PubMed] [Google Scholar]

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