Lymphocyte activation and capping of hormone receptors.
In this study both a ligand-dependent treatment [concanavalin A (Con A)] and a ligand-independent treatment [high-voltage pulsed galvanic stimulation (HVPGS)] have been used to initiate lymphocyte activation via a transmembrane signaling process. Our results show that both treatments cause the exposure of two different hormone [insulin and interleukin-2 (IL-2)] receptors within the first 5 min of stimulation. When either insulin or IL-2 is present in the culture medium, the stimulated lymphocytes undergo the following responses: (1) increased free intracellular Ca2+ activity; (2) aggregation of insulin or IL-2 receptors into patch/cap structures; (3) tyrosine-kinase-specific phosphorylation of a 32-kd membrane protein; and finally (4) induction of DNA synthesis. Further analysis indicates that hormone receptor capping is inhibited by (1) cytochalasin D, suggesting the involvement of microfilaments; (2) sodium azide, indicating a requirement for ATP production; and (3) W-5, W-7, and W-12 drugs, implying a need for Ca2+/calmodulin activity. Treatment with these metabolic or cytoskeletal inhibitors also prevents both the tyrosine-kinase-specific protein phosphorylation and DNA synthesis which normally follow hormone receptor capping. Double immunofluorescence staining shows that actomyosin, Ca2+/calmodulin, and myosin light-chain kinase are all closely associated with the insulin and IL-2 receptor cap structures. These findings strongly suggest that an actomyosin-mediated contractile system (regulated by Ca2+, calmodulin, and myosin light-chain kinase in an energy-dependent manner) is required not only for the collection of insulin and IL-2 receptors into patch and cap structures but also for the subsequent activation of tyrosine kinase and the initiation of DNA synthesis. We, therefore, propose that the exposure and subsequent patching/capping of at least one hormone receptor are required for the activation of mouse splenic T-lymphocytes.
D000818:Animals; D002118:Calcium / Q000378:metabolism; D003208:Concanavalin A / Q000494:pharmacology; D004247:DNA / Q000096:biosynthesis; D004558:Electric Stimulation; D007328:Insulin / Q000378:metabolism; D007376:Interleukin-2 / Q000378:metabolism; D008213:Lymphocyte Activation; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D010766:Phosphorylation; D011940:Receptor Aggregation; D011972:Receptor, Insulin / Q000378:metabolism*; D011971:Receptors, Immunologic / Q000378:metabolism*; D015375:Receptors, Interleukin-2; D013154:Spleen / Q000166:cytology; D013601:T-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D013997:Time Factors
D007328:Insulin; D007376:Interleukin-2; D008565:Membrane Proteins; D011971:Receptors, Immunologic; D015375:Receptors, Interleukin-2; D003208:Concanavalin A; D004247:DNA; D011972:Receptor, Insulin; D002118:Calcium