ALBUMIN ENDOCYTOSIS IN OK CELLS - DEPENDENCE ON ACTIN AND MICROTUBULES AND REGULATION BY PROTEIN-KINASES

We used proximal tubule-derived opossum kidney (OK) cells to determine the dependence of albumin endocytosis on regulation by protein kinase s and on the cytoskeleton. Uptake was observed only across the apical but not the basolateral membrane and exceeded uptake in collecting duc t-derived Madin-Darby canine kidney cells 14-fold. inhibition of endoc ytosis via clathrin-coated vesicles but not via caveolae abolished upt ake. Cytochalasin D reduced uptake to <5% of control, and inhibition o f microtubule polymerization by nocodazole reduced uptake to similar t o 55% of control. Activation of protein kinase A (PKA) by adenosine 3' ,5'-cyclic monophosphate, forskolin, or parathyroid hormone (PTH) redu ced uptake to similar to 65% of control. Protein kinase C (PKC) activa tion did affect uptake to a similar extent as PKA activation but with a certain delay Stimulation of PKG by guanosine 3',5'-cyclic monophosp hate did not affect albumin endocytosis. The inhibitor of tyrosine kin ases (TRK), genistein, induced an increase of uptake to similar to 160 % of control. Reexocytosis of albumin was enhanced by PKC activation b ut not by PKA activation. TRK inhibition reduced the rate of reexocyto sis. We conclude that albumin endocytosis in OK cells requires the int egrity of the actin cytoskeleton. Microtubules facilitate endocytosis. Uptake is regulated by PKA, PKC, and TRK, yet with different time cou rse and by different mechanisms, e.g., reexocytosis. Possibly TRK acti vity serves in a negative feedback loop to limit albumin endocytosis v ia a stimulation of reexocytosis.