Intramembrane Particles and the Organization of Lymphocyte Membrane Proteins

An experimental system was developed in which the majority of all lymphocyte cell-surface proteins, regardless of antigenic specificity, could be cross-linked and redistributed in the membrane to determine whether this would induce a corresponding redistribution of intramembrane particles (IMP) . Mouse spleen cells were treated with p-diazoniumphenyl-ßD-lactoside (lac) to modify all exposed cell-surface proteins . Extensive azo-coupling was achieved without significantly reducing cell viability or compromising cellular function in mitogenor antigen-stimulated cultures . When the Lac-modified cell-surface proteins were capped with a sandwich of rabbit antilactoside antibody and fluorescein-goat anti-rabbit Ig, freeze-fracture preparations obtained from these cells revealed no obvious redistribution of IMP on the majority of fracture faces. However, detailed analysis showed a statistically significant 35% decrease (P < 0.01) in average IMP density in the E face of the lac-capped spleen cells compared with control cells, whereas a few E-face micrographs showed intense IMP aggregation . In contrast, there was no significant alteration of P-face IMP densities or distribution . Apparently, the majority of E-face IMP and virtually all P-face IMP do not present accessible antigenic sites on the lymphocyte surface and do not associate in a stable manner with surface protein antigens . This finding suggests that IMP, as observed in freeze-fracture analysis, may not comprise a representative reflection of lymphocyte transmembrane protein molecules and complexes because other evidence establishes : (a) that at least some common lymphocyte surface antigens are indeed exposed portions of transmembrane proteins and (b) that the aggregation of molecules of any surface antigen results in altered organization of contractile proteins at the cytoplasmic face of the membrane . The role of the plasma membrane in the transmission of information can be seen in a number of systems where cells have been shown to be activated by an event at the cell membrane leading to proliferation and/or differentiation (5, 8, 23, 26) . Although the mechanism of lymphocyte activation is not known, there is evidence to suggest that the binding of insolubilized antigen or mitogen to membrane receptors may, under some conditions, be sufficient to initiate the cellular activation process (8) . There is considerable evidence for an association between lymphocyte cell-surface proteins and an underlying actinmyosin contractile system (2, 10, 15), which may be critically involved in the transmission of signals to or from the cell interior . However, the role of intramembrane particles (IMP) in this interaction remains unresolved. In erythrocytes, experiments have suggested an interaction of IMP with the underlying peripheral protein, spectrin, as well as with erythrocyte surface proteins (24) . In the lymphocyte membrane, associaTHE JOURNAL Of CELL BIOLOGY " VOLUME 88 MARCH 1981 591-598 ©The Rockefeller University Press " 0021-9525/81/03/0591/08 $1 .00 tions between IMP and either cell membrane receptors or an underlying contractile system have not been established . This research focuses on one aspect of the structure and organization of the lymphocyte plasma membrane by asking whether cross-linking and redistribution of lymphocyte surface proteins alter the distribution of IMP. A number of investigators have explored the consequence ofaggregating a portion of surface antigens with either antiimmunoglobuhns or lectins . These investigations have failed to detect or establish unambiguously the redistribution of IMP corresponding to the aggregation of the selected surface antigens (13, 16, 17), although the possibility always remains that, had redistribution of the right antigen been achieved, a corresponding redistribution of IMP might have been observed . To minimize this possibility, a system was devised where the majority of cell-surface proteins could be redistributed to determine whether there was a corresponding redistribution of IMP . Hapten groups were covalently coupled randomly to cell591 on Jne 8, 2017 D ow nladed fom Published March 1, 1981