Shear stress-dependent regulation of apical endocytosis in renal proximal tubule cells mediated by primary cilia

Abstract

The kidney has an extraordinary ability to maintain stable fractional solute and fluid reabsorption over a wide range of glomerular filtration rates (GFRs). Internalization of filtered low molecular weight proteins, vitamins, hormones, and other small molecules is mediated by the proximal tubule (PT) multiligand receptors megalin and cubilin. Changes in GFR and the accompanying fluid shear stress (FSS) modulate acute changes in PT ion transport thought to be mediated by microvillar bending. We found that FSS also affects apical endocytosis in PT cells. Exposure of immortalized PT cell lines to physiologically relevant levels of FSS led to dramatically increased internalization of the megalin-cubilin ligand albumin as well as the fluid phase marker dextran. FSS-stimulated apical endocytosis was initiated between 15 and 30 min postinduction of FSS, occurred via a clathrin- and dynamindependent pathway, and was rapidly reversed upon removing the FSS. Exposure to FSS also caused a rapid elevation in intracellular Ca 2+ [Ca 2+ ] i , which was not observed in deciliated cells, upon treatment with BAPTA-AM, or upon inclusion of apyrase in the perfusion medium. Strikingly, deciliation, BAPTA-AM, and apyrase also blocked the flow-dependent increase in endocytosis. Moreover, addition of ATP bypassed the need for FSS in enhancing endocytic capacity. Our studies suggest that increased [Ca 2+ ] i and purinergic signaling in response to FSS-dependent ciliary bending triggers a rapid and reversible increase in apical endocytosis that contributes to the efficient retrieval of filtered proteins in the PT.