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. 1985 Dec 1;101(6):2145–2155. doi: 10.1083/jcb.101.6.2145

Membrane and cytoskeletal changes associated with IgE-mediated serotonin release from rat basophilic leukemia cells

PMCID: PMC2113986  PMID: 2933414

Abstract

Binding of antigen to IgE-receptor complexes on the surface of RBL-2H3 rat basophilic leukemia cells is the first event leading to the release of cellular serotonin, histamine, and other mediators of allergic, asthmatic, and inflammatory responses. We have used dinitrophenol- conjugated bovine serum albumin (DNP-BSA) as well as the fluorescent antigen, DNP-B-phycoerythrin, and the electron-dense antigen, DNP-BSA- gold, to investigate dynamic membrane and cytoskeletal events associated with the release of [3H]serotonin from anti-DNP-IgE-primed RBL-2H3 cells. These multivalent antigens bind rapidly to cell surface IgE-receptor complexes. Their distribution is initially uniform, but within 2 min DNP-BSA-gold is found in coated pits and is subsequently internalized. Antigen internalization occurs in the presence and absence of extracellular Ca2+. The F-actin content of the detergent- extracted cell matrices analyzed by SDS PAGE decreases during the first 10-30 s of antigen binding and then increases by 1 min to almost double the control levels. A rapid and sustained increase is also observed when total F-actin is quantified by flow cytometry after binding of rhodamine-phalloidin. The antigen-stimulated increase in F-actin coincides with (and may cause) the transformation of the cell surface from a finely microvillous to a highly folded or plicated topography. Other early membrane responses include increased cell spreading and a 2- 3-fold increase in the uptake of fluorescein-dextran by fluid pinocytosis. The surface and F-actin changes show the same dependence on DNP-protein concentration as stimulated [3H]serotonin release; and both the membrane responses and the release of mediators are terminated by the addition of the non-cross-linking monovalent ligand, DNP-lysine. These data indicate that the same antigen-stimulated transduction pathway controls both the membrane/cytoskeletal and secretory events. However, the membrane and actin responses to IgE-receptor cross-linking are independent of extracellular Ca2+ and are mimicked by phorbol myristate acetate, whereas ligand-dependent mediator release depends on extracellular Ca2+ and is mimicked by the Ca2+ ionophore A23187.

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Selected References

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  1. Beaven M. A., Moore J. P., Smith G. A., Hesketh T. R., Metcalfe J. C. The calcium signal and phosphatidylinositol breakdown in 2H3 cells. J Biol Chem. 1984 Jun 10;259(11):7137–7142. [PubMed] [Google Scholar]
  2. Beaven M. A., Rogers J., Moore J. P., Hesketh T. R., Smith G. A., Metcalfe J. C. The mechanism of the calcium signal and correlation with histamine release in 2H3 cells. J Biol Chem. 1984 Jun 10;259(11):7129–7136. [PubMed] [Google Scholar]
  3. Becker E. L. The relationship of the chemotactic behavior of the complement-derived factors, C3a, C5a, and C567, and a bacterial chemotactic factor to their ability to activate the proesterase 1 of rabbit polymorphonuclear leukocytes. J Exp Med. 1972 Feb 1;135(2):376–387. doi: 10.1084/jem.135.2.376. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Braun J., Hochman P. S., Unanue E. R. Ligand-induced association of surface immunoglobulin with the detergent-insoluble cytoskeletal matrix of the B lymphocyte. J Immunol. 1982 Mar;128(3):1198–1204. [PubMed] [Google Scholar]
  5. Carson D. A., Metzger H. Interaction of IgE with rat basophilic leukemia cells. IV. Antibody-induced redistribution of IgE receptors. J Immunol. 1974 Oct;113(4):1271–1277. [PubMed] [Google Scholar]
  6. Chinkers M., McKanna J. A., Cohen S. Rapid induction of morphological changes in human carcinoma cells A-431 by epidermal growth factors. J Cell Biol. 1979 Oct;83(1):260–265. doi: 10.1083/jcb.83.1.260. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cockcroft S., Gomperts B. D. Role of guanine nucleotide binding protein in the activation of polyphosphoinositide phosphodiesterase. Nature. 1985 Apr 11;314(6011):534–536. doi: 10.1038/314534a0. [DOI] [PubMed] [Google Scholar]
  8. Condeelis J. Isolation of concanavalin A caps during various stages of formation and their association with actin and myosin. J Cell Biol. 1979 Mar;80(3):751–758. doi: 10.1083/jcb.80.3.751. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Connolly J. L., Green S. A., Greene L. A. Pit formation and rapid changes in surface morphology of sympathetic neurons in response to nerve growth factor. J Cell Biol. 1981 Jul;90(1):176–180. doi: 10.1083/jcb.90.1.176. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Davis B. H., Walter R. J., Pearson C. B., Becker E. L., Oliver J. M. Membrane activity and topography of F-Met-Leu-Phe-Treated polymorphonuclear leukocytes. Acute and sustained responses to chemotactic peptide. Am J Pathol. 1982 Aug;108(2):206–216. [PMC free article] [PubMed] [Google Scholar]
  11. Dvorak A. M., Galli S. J., Schulman E. S., Lichtenstein L. M., Dvorak H. F. Basophil and mast cell degranulation: ultrastructural analysis of mechanisms of mediator release. Fed Proc. 1983 May 15;42(8):2510–2515. [PubMed] [Google Scholar]
  12. Fewtrell C., Lagunoff D., Metzger H. Secretion from rat basophilic leukaemia cells induced by calcium ionophores. Effect of pH and metabolic inhibition. Biochim Biophys Acta. 1981 Jun 22;644(2):363–368. doi: 10.1016/0005-2736(81)90394-1. [DOI] [PubMed] [Google Scholar]
  13. Fewtrell C., Metzger H. Larger oligomers of IgE are more effective than dimers in stimulating rat basophilic leukemia cells. J Immunol. 1980 Aug;125(2):701–710. [PubMed] [Google Scholar]
  14. Furuichi K., Rivera J., Isersky C. The fate of IgE bound to rat basophilic leukemia cells. III. Relationship between antigen-induced endocytosis and serotonin release. J Immunol. 1984 Sep;133(3):1513–1520. [PubMed] [Google Scholar]
  15. Goshima K., Masuda A., Owaribe K. Insulin-induced formation of ruffling membranes of KB cells and its correlation with enhancement of amino acid transport. J Cell Biol. 1984 Mar;98(3):801–809. doi: 10.1083/jcb.98.3.801. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Heiman A. S., Crews F. T. Characterization of the effects of phorbol esters on rat mast cell secretion. J Immunol. 1985 Jan;134(1):548–555. [PubMed] [Google Scholar]
  17. Holowka D., Hartmann H., Kanellopoulos J., Metzger H. Association of the receptor for immunoglobulin E with an endogenous polypeptide on rat basophilic leukemia cells. J Recept Res. 1980;1(1):41–68. doi: 10.3109/10799898009039254. [DOI] [PubMed] [Google Scholar]
  18. Howard T. H., Meyer W. H. Chemotactic peptide modulation of actin assembly and locomotion in neutrophils. J Cell Biol. 1984 Apr;98(4):1265–1271. doi: 10.1083/jcb.98.4.1265. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Isersky C., Rivera J., Segal D. M., Triche T. The fate of IgE bound to rat basophilic leukemia cells. II. Endocytosis of IgE oligomers and effect on receptor turnover. J Immunol. 1983 Jul;131(1):388–396. [PubMed] [Google Scholar]
  20. Jesaitis A. J., Naemura J. R., Sklar L. A., Cochrane C. G., Painter R. G. Rapid modulation of N-formyl chemotactic peptide receptors on the surface of human granulocytes: formation of high-affinity ligand-receptor complexes in transient association with cytoskeleton. J Cell Biol. 1984 Apr;98(4):1378–1387. doi: 10.1083/jcb.98.4.1378. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kanner B. I., Metzger H. Crosslinking of the receptors for immunoglobulin E depolarizes the plasma membrane of rat basophilic leukemia cells. Proc Natl Acad Sci U S A. 1983 Sep;80(18):5744–5748. doi: 10.1073/pnas.80.18.5744. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  23. Liu F. T., Bohn J. W., Ferry E. L., Yamamoto H., Molinaro C. A., Sherman L. A., Klinman N. R., Katz D. H. Monoclonal dinitrophenyl-specific murine IgE antibody: preparation, isolation, and characterization. J Immunol. 1980 Jun;124(6):2728–2737. [PubMed] [Google Scholar]
  24. Martin J. C., Swartzendruber D. E. Time: a new parameter for kinetic measurements in flow cytometry. Science. 1980 Jan 11;207(4427):199–201. doi: 10.1126/science.6153131. [DOI] [PubMed] [Google Scholar]
  25. McCloskey M. A., Liu Z. Y., Poo M. M. Lateral electromigration and diffusion of Fc epsilon receptors on rat basophilic leukemia cells: effects of IgE binding. J Cell Biol. 1984 Sep;99(3):778–787. doi: 10.1083/jcb.99.3.778. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Mendoza G., Metzger H. Distribution and valency of receptor for IgE on rodent mast cells and related tumour cells. Nature. 1976 Dec 9;264(5586):548–550. doi: 10.1038/264548a0. [DOI] [PubMed] [Google Scholar]
  27. Menon A. K., Holowka D., Baird B. Small oligomers of immunoglobulin E (IgE) cause large-scale clustering of IgE receptors on the surface of rat basophilic leukemia cells. J Cell Biol. 1984 Feb;98(2):577–583. doi: 10.1083/jcb.98.2.577. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Nishizuka Y. The role of protein kinase C in cell surface signal transduction and tumour promotion. Nature. 1984 Apr 19;308(5961):693–698. doi: 10.1038/308693a0. [DOI] [PubMed] [Google Scholar]
  29. Oliver J. M., Seagrave J. C., Pfeiffer J. R., Feibig M. L., Deanin G. G. Surface functions during mitosis in rat basophilic leukemia cells. J Cell Biol. 1985 Dec;101(6):2156–2166. doi: 10.1083/jcb.101.6.2156. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Phillips D. R., Jennings L. K., Edwards H. H. Identification of membrane proteins mediating the interaction of human platelets. J Cell Biol. 1980 Jul;86(1):77–86. doi: 10.1083/jcb.86.1.77. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Rivnay B., Wank S. A., Poy G., Metzger H. Phospholipids stabilize the interaction between the alpha and beta subunits of the solubilized receptor for immunoglobulin E. Biochemistry. 1982 Dec 21;21(26):6922–6927. doi: 10.1021/bi00269a047. [DOI] [PubMed] [Google Scholar]
  32. Salzman G. C., Wilkins S. F., Whitfill J. A. Modular computer programs for flow cytometry and sorting: the LACEL system. Cytometry. 1981 Mar;1(5):325–336. doi: 10.1002/cyto.990010505. [DOI] [PubMed] [Google Scholar]
  33. Shechter Y., Chang K. J., Jacobs S., Cuatrecasas P. Modulation of binding and bioactivity of insulin by anti-insulin antibody: relation to possible role of receptor self-aggregation in hormone action. Proc Natl Acad Sci U S A. 1979 Jun;76(6):2720–2724. doi: 10.1073/pnas.76.6.2720. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Sheterline P., Hopkins C. R. Transmembrane linkage between surface glycoproteins and components of the cytoplasm in neutrophil leukocytes. J Cell Biol. 1981 Sep;90(3):743–754. doi: 10.1083/jcb.90.3.743. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Siraganian R. P., McGivney A., Barsumian E. L., Crews F. T., Hirata F., Axelrod J. Variants of the rat basophilic leukemia cell line for the study of histamine release. Fed Proc. 1982 Jan;41(1):30–34. [PubMed] [Google Scholar]
  36. Taurog J. D., Mendoza G. R., Hook W. A., Siraganian R. P., Metzger H. Noncytotoxic IgE-mediated release of histamine and serotonin from murine mastocytoma cells. J Immunol. 1977 Nov;119(5):1757–1761. [PubMed] [Google Scholar]
  37. Vale R. D., Shooter E. M. Alteration of binding properties and cytoskeletal attachment of nerve growth factor receptors in PC12 cells by wheat germ agglutinin. J Cell Biol. 1982 Sep;94(3):710–717. doi: 10.1083/jcb.94.3.710. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Wallace P. J., Wersto R. P., Packman C. H., Lichtman M. A. Chemotactic peptide-induced changes in neutrophil actin conformation. J Cell Biol. 1984 Sep;99(3):1060–1065. doi: 10.1083/jcb.99.3.1060. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Woda B. A., Woodin M. B. The interaction of lymphocyte membrane proteins with the lymphocyte cytoskeletal matrix. J Immunol. 1984 Nov;133(5):2767–2772. [PubMed] [Google Scholar]

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