Evidence of a structural and functional ammonium transporter RhBG·anion exchanger 1·ankyrin-G complex in kidney epithelial cells.

摘要: The renal ammonium transporter RhBG and anion exchanger 1 kAE1 colocalize in the basolateral domain of α-intercalated cells distal nephron. Although we have previously shown that is linked to spectrin-based skeleton through ankyrin-G its NH3 transport activity dependent on this association, there no evidence for an interaction with adaptor protein. We report here cytoplasmic N terminus actually binds ankyrin-G, both yeast two-hybrid analysis by coimmunoprecipitation situ HEK293 expressing recombinant kAE1. A site-directed mutagenesis study allowed identification three dispersed regions molecule linking third fourth repeat domains ankyrin-G. One secondary docking site corresponds a major interacting loop erythroid (eAE1) ankyrin-R, whereas main binding region does not encompass any eAE1 determinant. Stopped flow spectrofluorometry revealed Cl−/HCO3− exchange protein mutated was abolished. This disruption impaired plasma membrane expression leading total retention structures polarized epithelial Madin-Darby canine kidney cell transfectants. also directly interacts without affecting surface function. first description structural functional RhBG·kAE1·ankyrin-G complex at cells, comparable well known Rh·eAE1·ankyrin-R red blood membrane. could participate regulation acid-base homeostasis.