THE EFFECTS OF CYTOCHALASIN-D AND PHORBOL-MYRISTATE ACETATE ON THE APICAL ENDOCYTOSIS OF RICIN IN POLARIZED CACO-2 CELLS

Apical endocytosis of I-125-ricin in Caco-2 cells was inhibited >95% b y hypertonic and/or acid media, consistent with the major uptake route being clathrin-mediated. The presence of apical cell surface bound ri cin-gold in clathrin coated pits and vesicles was observed by electron microscopy. An electron microscopic investigation in which ricin-gold bound to the apical surface was quantitated, showed that cytochalasin D, which inhibits apical but not basolateral endocytosis, prevented m ovement of ricin-gold along the microvillar surface. This was consiste nt with an actin bound mechanochemical motor within microvilli driving the movement of membranous components towards the cell body. Cytochal asin D also caused an increase in the number of coated pits observed a t the apical cell surface relative to the number observed in untreated cells, Stimulation of apical endocytosis of ricin by phorbol 12-myris tate 13-acetate showed the characteristics of being mediated by protei n kinase C, was not due to an effect on ricin movement along the micro villar surface, and may be explained by increases in formation and pin ching off of clathrin coated pits at the apical cell surface.