| Abstract: | Protein kinase C (PKC) and the actin cytoskeleton are criticaleffectors of membrane trafficking in mammalian cells. In polarized epithelia, the role of these factors in endocytic events at either theapical or basolateral membrane is poorly defined. In the present study,phorbol 12-myristate 13-acetate (PMA) and other activators of PKCselectively enhanced basolateral but not apical fluid-phase endocytosisin human T84 intestinal epithelia. Stimulation of basolateralendocytosis was blocked by the conventional and novel PKC inhibitorGö-6850, but not the conventional PKC inhibitor Gö-6976,and correlated with translocation of the novel PKC isoform PKC- . PMAtreatment induced remodeling of basolateral F-actin. The actindisassembler cytochalasin D stimulated basolateral endocytosis andenhanced stimulation of endocytosis by PMA, whereas PMA-stimulated endocytosis was blocked by the F-actin stabilizers phalloidin andjasplakinolide. PMA induced membrane-to-cytosol redistribution of theF-actin cross-linking protein myristoylated alanine-rich C kinasesubstrate (MARCKS). Cytochalasin D also induced MARCKS translocationand enhanced PMA-stimulated translocation of MARCKS. A myristoylatedpeptide corresponding to the phosphorylation site domain of MARCKSinhibited both MARCKS translocation and PMA stimulation of endocytosis.MARCKS translocation was inhibited by Gö-6850 but notGö-6976. The results suggest that a novel PKC isoform, likelyPKC-, stimulates basolateral endocytosis in model epithelia by amechanism that involves F-actin and MARCKS. |
