Receptor capping in mouse T-lymphoma cells: a Ca2+ and calmodulin-stimulated ATP-dependent process.
The roles that Ca2+, calmodulin, and ATP play in the redistribution of concanavalin A (Con A) binding sites on the surface of mouse T-lymphoma cells were examined. Membranes of cells labeled with fluorescein-conjugated Con A (Fl-Con A) were made permeable ("skinned") to ions and proteins by incubation in a solution containing no added Ca2+, 7 mM EGTA, and ATP. The intracellular ionic and protein concentrations could then be varied, and the degree of Con A receptor capping monitored simultaneously. A graded increase (9.0 to 30%) was found in the number of capped cells with increasing Ca2+ concentration from 10(-6)-10(-4.9) M. Increasing concentrations of trifluoperazine, chlorpromazine, and promethazine (1.5 x 10(-6) to 1.0 x 10(-4) M) in cell suspensions containing 10(-4) M Ca2+ produced graded inhibition of capping in the same order that the drugs bind to calmodulin. Removal of extracellular Ca2+ dissociated (reversed) some of the caps into patches, thus reducing their number (12%). ATP was required for either capping or cap dissociation to occur. Addition of calmodulin (3.9 x 10(-8)-6.3 x 10(-7) M) to the cell suspension increased the Ca2+ sensitivity. These results provide direct evidence that capping of Con A receptors is a reversible process (i) regulated by intracellular Ca2+ concentrations, (ii) requiring ATP as an energy source, and (iii) susceptible to the influence of calmodulin. These findings are consistent with the hypothesis that the collection of surface receptor patches into cap structures is controlled by the interaction of actomyosin filaments, which in turn is regulated by a Ca2+-calmodulin-activated control system.
D000255:Adenosine Triphosphate / Q000502:physiology; D000818:Animals; D002118:Calcium / Q000502:physiology*; D002135:Calcium-Binding Proteins / Q000502:physiology*; D002147:Calmodulin / Q000502:physiology*; D002463:Cell Membrane Permeability; D002478:Cells, Cultured; D007152:Immunologic Capping; D008223:Lymphoma / Q000276:immunology; D051379:Mice; D013601:T-Lymphocytes / Q000276:immunology*
D002135:Calcium-Binding Proteins; D002147:Calmodulin; D000255:Adenosine Triphosphate; D002118:Calcium