Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1987 Sep 1;105(3):1147–1152. doi: 10.1083/jcb.105.3.1147

Lateral mobility of class I histocompatibility antigens in B lymphoblastoid cell membranes: modulation by cross-linking and effect of cell density

PMCID: PMC2114826  PMID: 3115995

Abstract

We have studied the lateral mobility of class 1 major histocompatibility complex (MHC) proteins in the membranes of human Epstein-Barr virus-transformed B cells using fluorescence photobleaching recovery. Class I MHC antigens were labeled with either W6/32 monoclonal antibody or its Fab fragment directly conjugated to fluorescein isothiocyanate. The diffusion coefficient of class I antigens labeled with Fab fragments of W6/32 was identical to that of a lipid analogue, fluorescein phosphatidylethanolamine, and was 10-fold greater than that of antigens labeled with intact W6/32. Furthermore, antigens labeled with Fab fragments but not with intact W6/32 had fractional mobilities identical to that of the lipid probe. The lateral mobility of class I antigens was dependent on the time of incubation with fluorescent antibody and on the presence of antibody microaggregates. Finally, class I MHC proteins labeled with intact W6/32 but not with Fab fragments were immobilized in the membranes of most cells grown in suspension at high cell density. These results suggest that, in the unperturbed state, class I MHC antigens diffuse as rapidly as membrane lipid, i.e., without cytoskeletal constraint. Cross- linking with bivalent ligand and growth to high cell density may trigger membrane events leading to slowing and immobilization of these proteins.

Full Text

The Full Text of this article is available as a PDF (804.6 KB).

Selected References

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

  1. Axelrod D., Koppel D. E., Schlessinger J., Elson E., Webb W. W. Mobility measurement by analysis of fluorescence photobleaching recovery kinetics. Biophys J. 1976 Sep;16(9):1055–1069. doi: 10.1016/S0006-3495(76)85755-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Axelrod D. Lateral motion of membrane proteins and biological function. J Membr Biol. 1983;75(1):1–10. doi: 10.1007/BF01870794. [DOI] [PubMed] [Google Scholar]
  3. Bloom J. A., Webb W. W. Lipid diffusibility in the intact erythrocyte membrane. Biophys J. 1983 Jun;42(3):295–305. doi: 10.1016/S0006-3495(83)84397-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Braun J., Fujiwara K., Pollard T. D., Unanue E. R. Two distinct mechanisms for redistribution of lymphocyte surface macromolecules. I. Relationship to cytoplasmic myosin. J Cell Biol. 1978 Nov;79(2 Pt 1):409–418. doi: 10.1083/jcb.79.2.409. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. 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]
  6. Damjanovich S., Trón L., Szöllösi J., Zidovetzki R., Vaz W. L., Regateiro F., Arndt-Jovin D. J., Jovin T. M. Distribution and mobility of murine histocompatibility H-2Kk antigen in the cytoplasmic membrane. Proc Natl Acad Sci U S A. 1983 Oct;80(19):5985–5989. doi: 10.1073/pnas.80.19.5985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Dragsten P., Henkart P., Blumenthal R., Weinstein J., Schlessinger J. Lateral diffusion of surface immunoglobulin, Thy-1 antigen, and a lipid probe in lymphocyte plasma membranes. Proc Natl Acad Sci U S A. 1979 Oct;76(10):5163–5167. doi: 10.1073/pnas.76.10.5163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Edidin M., Wei T. Lateral diffusion of H-2 antigens on mouse fibroblasts. J Cell Biol. 1982 Nov;95(2 Pt 1):458–462. doi: 10.1083/jcb.95.2.458. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Edidin M., Zuniga M. Lateral diffusion of wild-type and mutant Ld antigens in L cells. J Cell Biol. 1984 Dec;99(6):2333–2335. doi: 10.1083/jcb.99.6.2333. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Fehlmann M., Peyron J. F., Samson M., Van Obberghen E., Brandenburg D., Brossette N. Molecular association between major histocompatibility complex class I antigens and insulin receptors in mouse liver membranes. Proc Natl Acad Sci U S A. 1985 Dec;82(24):8634–8637. doi: 10.1073/pnas.82.24.8634. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Galla H. J., Hartmann W., Theilen U., Sackmann E. On two-dimensional passive random walk in lipid bilayers and fluid pathways in biomembranes. J Membr Biol. 1979 Jul 31;48(3):215–236. doi: 10.1007/BF01872892. [DOI] [PubMed] [Google Scholar]
  12. Golan D. E., Brown C. S., Cianci C. M., Furlong S. T., Caulfield J. P. Schistosomula of Schistosoma mansoni use lysophosphatidylcholine to lyse adherent human red blood cells and immobilize red cell membrane components. J Cell Biol. 1986 Sep;103(3):819–828. doi: 10.1083/jcb.103.3.819. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Henis Y. I., Gutman O. Lateral diffusion and patch formation of H-2Kk antigens on mouse spleen lymphocytes. Biochim Biophys Acta. 1983 Apr 5;762(2):281–288. doi: 10.1016/0167-4889(83)90082-4. [DOI] [PubMed] [Google Scholar]
  14. Kapitza H. G., Sackmann E. Local measurement of lateral motion in erythrocyte membranes by photobleaching technique. Biochim Biophys Acta. 1980;595(1):56–64. doi: 10.1016/0005-2736(80)90247-3. [DOI] [PubMed] [Google Scholar]
  15. Koch G. L., Smith M. J. An association between actin and the major histocompatibility antigen H-2. Nature. 1978 May 25;273(5660):274–278. doi: 10.1038/273274a0. [DOI] [PubMed] [Google Scholar]
  16. Krangel M. S., Orr H. T., Strominger J. L. Structure, function, and biosynthesis of the major human histocompatibility antigens (HLA-A and HLA-B). Scand J Immunol. 1980;11(6):561–571. doi: 10.1111/j.1365-3083.1980.tb00025.x. [DOI] [PubMed] [Google Scholar]
  17. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  18. 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]
  19. Menon A. K., Holowka D., Webb W. W., Baird B. Clustering, mobility, and triggering activity of small oligomers of immunoglobulin E on rat basophilic leukemia cells. J Cell Biol. 1986 Feb;102(2):534–540. doi: 10.1083/jcb.102.2.534. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Menon A. K., Holowka D., Webb W. W., Baird B. Cross-linking of receptor-bound IgE to aggregates larger than dimers leads to rapid immobilization. J Cell Biol. 1986 Feb;102(2):541–550. doi: 10.1083/jcb.102.2.541. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Merril C. R., Goldman D., Sedman S. A., Ebert M. H. Ultrasensitive stain for proteins in polyacrylamide gels shows regional variation in cerebrospinal fluid proteins. Science. 1981 Mar 27;211(4489):1437–1438. doi: 10.1126/science.6162199. [DOI] [PubMed] [Google Scholar]
  22. Nakache M., Schreiber A. B., Gaub H., McConnell H. M. Heterogeneity of membrane phospholipid mobility in endothelial cells depends on cell substrate. Nature. 1985 Sep 5;317(6032):75–77. doi: 10.1038/317075a0. [DOI] [PubMed] [Google Scholar]
  23. Parham P., Barnstable C. J., Bodmer W. F. Use of a monoclonal antibody (W6/32) in structural studies of HLA-A,B,C, antigens. J Immunol. 1979 Jul;123(1):342–349. [PubMed] [Google Scholar]
  24. Petty H. R., Smith L. M., Fearon D. T., McConnell H. M. Lateral distribution and diffusion of the C3b receptor of complement, HLA antigens, and lipid probes in peripheral blood leukocytes. Proc Natl Acad Sci U S A. 1980 Nov;77(11):6587–6591. doi: 10.1073/pnas.77.11.6587. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Phillips M. L., Moule M. L., Delovitch T. L., Yip C. C. Class I histocompatibility antigens and insulin receptors: evidence for interactions. Proc Natl Acad Sci U S A. 1986 May;83(10):3474–3478. doi: 10.1073/pnas.83.10.3474. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Pober J. S., Guild B. C., Strominger J. L., Veatch W. R. Purification of HLA-A2 antigen, fluorescent labeling of its intracellular region, and demonstration of an interaction between fluorescently labeled HLA-A2 antigen and lymphoblastoid cell cytoskeleton proteins in vitro. Biochemistry. 1981 Sep 15;20(19):5625–5633. doi: 10.1021/bi00522a042. [DOI] [PubMed] [Google Scholar]
  27. Saffman P. G., Delbrück M. Brownian motion in biological membranes. Proc Natl Acad Sci U S A. 1975 Aug;72(8):3111–3113. doi: 10.1073/pnas.72.8.3111. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Schreiber A. B., Schlessinger J., Edidin M. Interaction between major histocompatibility complex antigens and epidermal growth factor receptors on human cells. J Cell Biol. 1984 Feb;98(2):725–731. doi: 10.1083/jcb.98.2.725. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Schwartz R. H. T-lymphocyte recognition of antigen in association with gene products of the major histocompatibility complex. Annu Rev Immunol. 1985;3:237–261. doi: 10.1146/annurev.iy.03.040185.001321. [DOI] [PubMed] [Google Scholar]
  30. Smith L. M., Petty H. R., Parham P., McConnell H. M. Cell surface properties of HLA antigens on Epstein-Barr virus-transformed cell lines. Proc Natl Acad Sci U S A. 1982 Jan;79(2):608–612. doi: 10.1073/pnas.79.2.608. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Smith L. M., Rubenstein J. L., Parce J. W., McConnell H. M. Lateral diffusion of M-13 coat protein in mixtures of phosphatidylcholine and cholesterol. Biochemistry. 1980 Dec 9;19(25):5907–5911. doi: 10.1021/bi00566a037. [DOI] [PubMed] [Google Scholar]
  32. Su Y. X., Lin S., Edidin M. Lateral diffusion of human histocompatibility antigens in isolated plasma membranes. Biochim Biophys Acta. 1984 Sep 19;776(1):92–96. doi: 10.1016/0005-2736(84)90254-2. [DOI] [PubMed] [Google Scholar]
  33. Subramaniam S., Seul M., McConnell H. M. Lateral diffusion of specific antibodies bound to lipid monolayers on alkylated substrates. Proc Natl Acad Sci U S A. 1986 Mar;83(5):1169–1173. doi: 10.1073/pnas.83.5.1169. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Tse D. B., Pernis B. Spontaneous internalization of Class I major histocompatibility complex molecules in T lymphoid cells. J Exp Med. 1984 Jan 1;159(1):193–207. doi: 10.1084/jem.159.1.193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Vaz W. L., Criado M., Madeira V. M., Schoellmann G., Jovin T. M. Size dependence of the translational diffusion of large integral membrane proteins in liquid-crystalline phase lipid bilayers. A study using fluorescence recovery after photobleaching. Biochemistry. 1982 Oct 26;21(22):5608–5612. doi: 10.1021/bi00265a034. [DOI] [PubMed] [Google Scholar]
  36. Wier M. L., Edidin M. Effects of cell density and extracellular matrix on the lateral diffusion of major histocompatibility antigens in cultured fibroblasts. J Cell Biol. 1986 Jul;103(1):215–222. doi: 10.1083/jcb.103.1.215. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Woda B. A., Gilman S. C. Lateral mobility and capping of rat lymphocyte membrane proteins. Cell Biol Int Rep. 1983 Mar;7(3):203–209. doi: 10.1016/0309-1651(83)90227-8. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES