Requirement of cortical actin organization for bombesin, endothelin, and EGF receptor internalization

The role of actin organization in occupancy-induced receptor internalizatio n remains poorly defined. Here we report that treatment of mouse Swiss 3T3 cells with latrunculin A, a potent inhibitor of actin polymerization (inclu ding cortical actin), inhibited the internalization of the endogenous bombe sin/gastrin-releasing peptide (GRP) receptor, as judged by uptake of I-125- labeled GRP or fluorescent Cy3-labeled bombesin. In contrast, cells pretrea ted with cytochalasin D showed minimal inhibition of bombesin/GRP receptor internalization. Similarly, pretreatment of Swiss 3T3 cells with the potent Rho-kinase inhibitor HA-1077, at concentrations (10-20 muM) that abrogated bombesin-mediated stress fiber formation, did not significantly alter rece ptor-mediated internalization of I-125-GRP. These results indicate that bom besin/GRP receptor internalization depends on latrunculin A-sensitive corti cal actin rather than on rapidly turning over actin stress fibers that are disrupted by either cytochalasin D or HA-1077. The rates and total levels o f internalization of the endogenously expressed endothelin A receptor and e pidermal growth factor receptor were also markedly reduced by latrunculin A in Swiss 3T3 cells. The potency of latrunculin A for inhibiting G protein- coupled receptor endocytosis was comparable to that for reducing internaliz ation of the epidermal growth factor tyrosine kinase receptor. We conclude that cortical actin structures, disrupted by latrunculin A, are necessary f or occupancy-induced receptor internalization in animal cells.