Opposing effects of PKCA andpkce on basolateral membrane dynamics in intestinal epithelia

Abstract

Song, Jaekyung Cecilia, Patangi K. Rangachari, and Jeffrey B. Matthews. Opposing effects of PKC and PKC on basolateral membrane dynamics in intestinal epithelia. Am J Physiol Cell Physiol 283: C1548–C1556, 2002. First published July 24, 2002; 10.1152/ajpcell.00105.2002.—PKC is a critical effector of plasma membrane dynamics, yet the mechanism and isoform-specific role of PKC are poorly understood. We recently showed that the phorbol ester PMA (100 nM) induces prompt activation of the novel isoform PKC followed by late activation of the conventional isoform PKC in T84 intestinal epithelia. PMA also elicited biphasic effects on endocytosis, characterized by an initial stimulatory phase followed by an inhibitory phase. Activation of PKC was shown to be responsible for stimulation of basolateral endocytosis, but the role of PKC was not defined. Here, we used detailed time-course analysis as well as selective activators and inhibitors of PKC isoforms to infer the action of PKC on basolateral endocytosis. Inhibition of PKC by the selective conventional PKC inhibitor Go-6976 (5 M) completely blocked the late inhibitory phase and markedly prolonged the stimulatory phase of endocytosis measured by FITC-dextran uptake. The PKC -selective agonist carbachol (100 M) induced prolonged stimulation of endocytosis devoid of an inhibitory phase. Actin disassembly caused by PMA was completely blocked by Go-6850 but not by Go-6976, implicating PKC as the key isoform responsible for actin disruption. The Ca2 agonist thapsigargin (5 M) induced early activation of PKC when added simultaneously with PMA. This early activation of PKC blocked the ability of PMA to remodel basolateral F-actin and abolished the stimulatory phase of basolateral endocytosis. Activation of PKC stabilizes F-actin and thereby opposes the effect of PKC on membrane remodeling in T84 cells.