Abstract
We have used fluorescence photobleaching and recovery (FPR) to measure the lateral diffusion of mouse H-2 antigens, labeled with fluorescent Fab fragments, in the membrane of cl 1d fibroblasts. Diffusion coefficients, D, vary more than 20-fold from cell to cell, though they vary no more than twofold when measured at different points on a single cell. The fraction of H-2 antigens mobile, R, also varies from cell to cell, and no lateral diffusion of H-2 antigens can be detected in approximately 20% of the cells examined. Treatment of cells with NaCN + NaF, reducing their levels of ATP reduces the proportion of cells in which no lateral diffusion can be detected. The maximum values of D seen in poisoned cells are less than those in controls. Treatment of cells with the divalent inophore, A23187, greatly increases the proportion of cells in which diffusion of H-2 is rapid, D greater than 2 x 10(-9) cm2 s-1. The data obtained on diffusion by FPR can be replotted in the form of an experiment in which lateral diffusion of H- 2 antigens is measured in a population of heterokaryons. There is good agreement between this transformation and actual data on heterokaryons. Thus the two methods appear to measure the same transport process.
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- Axelrod D. Cell surface heating during fluorescence photobleaching recovery experiments. Biophys J. 1977 Apr;18(1):129–131. doi: 10.1016/S0006-3495(77)85601-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- Barisas B. G., Leuther M. D. Fluorescence photobleaching recovery measurement of protein absolute diffusion constants. Biophys Chem. 1979 Sep;10(2):221–229. doi: 10.1016/0301-4622(79)85044-9. [DOI] [PubMed] [Google Scholar]
- Bloch R. J. Dispersal and reformation of acetylcholine receptor clusters of cultured rat myotubes treated with inhibitors of energy metabolism. J Cell Biol. 1979 Sep;82(3):626–643. doi: 10.1083/jcb.82.3.626. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cebra J. J., Goldstein G. Chromatographic purification of tetramethylrhodamine-immune globulin conjugates and their use in the cellular localization of rabbit gamma-globulin polypeptide chains. J Immunol. 1965 Aug;95(2):230–245. [PubMed] [Google Scholar]
- Edidin M., Wei T. Y. Diffusion rates of cell surface antigens of mouse-human heterokaryons. I. Analysis of the population. J Cell Biol. 1977 Nov;75(2 Pt 1):475–482. doi: 10.1083/jcb.75.2.475. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Edidin M., Wei T. Y. Diffusion rates of cell surface antigens of mouse-human heterokaryons. II. Effect of membrane potential on lateral diffusion. J Cell Biol. 1977 Nov;75(2 Pt 1):483–489. doi: 10.1083/jcb.75.2.483. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Edidin M., Wei T., Gotlib L. Diffusion rates of cell surface antigens of mouse-human heterokaryons. III. Regulation of lateral diffusion rates by calcium ions. J Cell Biol. 1982 Nov;95(2 Pt 1):453–462. doi: 10.1083/jcb.95.2.453. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ey P. L., Prowse S. J., Jenkin C. R. Isolation of pure IgG1, IgG2a and IgG2b immunoglobulins from mouse serum using protein A-sepharose. Immunochemistry. 1978 Jul;15(7):429–436. doi: 10.1016/0161-5890(78)90070-6. [DOI] [PubMed] [Google Scholar]
- Frye L. D., Edidin M. The rapid intermixing of cell surface antigens after formation of mouse-human heterokaryons. J Cell Sci. 1970 Sep;7(2):319–335. doi: 10.1242/jcs.7.2.319. [DOI] [PubMed] [Google Scholar]
- Golan D. E., Veatch W. Lateral mobility of band 3 in the human erythrocyte membrane studied by fluorescence photobleaching recovery: evidence for control by cytoskeletal interactions. Proc Natl Acad Sci U S A. 1980 May;77(5):2537–2541. doi: 10.1073/pnas.77.5.2537. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Guyer R. L., Koshland M. E., Knopf P. M. Immunoglobulin binding by mouse intestinal epithelial cell receptors. J Immunol. 1976 Aug;117(2):587–593. [PubMed] [Google Scholar]
- Koppel D. E. Fluorescence redistribution after photobleaching. A new multipoint analysis of membrane translational dynamics. Biophys J. 1979 Nov;28(2):281–291. doi: 10.1016/S0006-3495(79)85176-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Oi V. T., Jones P. P., Goding J. W., Herzenberg L. A., Herzenberg L. A. Properties of monoclonal antibodies to mouse Ig allotypes, H-2, and Ia antigens. Curr Top Microbiol Immunol. 1978;81:115–120. doi: 10.1007/978-3-642-67448-8_18. [DOI] [PubMed] [Google Scholar]
- Ozato K., Mayer N., Sachs D. H. Hybridoma cell lines secreting monoclonal antibodies to mouse H-2 and Ia antigens. J Immunol. 1980 Feb;124(2):533–540. [PubMed] [Google Scholar]
- 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]
- Reed P. W., Lardy H. A. A23187: a divalent cation ionophore. J Biol Chem. 1972 Nov 10;247(21):6970–6977. [PubMed] [Google Scholar]
- 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]