An Essential Role of Nedd4-2 in the Ubiquitination, Expression, and Function of Organic Anion Transporter-3.

Organic anion transporter-3 (OAT3) is a member of the organic anion transporter family that mediates the body disposition of a diverse array of clinically important drugs. We previously demonstrated that activation of protein kinase C (PKC) inhibits OAT3 transport activity by accelerating OAT3 internalization from cell surface into intracellular compartments. In the current study, we established that PKC-induced inhibition of OAT3 transport activity occurred through an enhanced OAT3 ubiquitination, a process catalyzed by an E3 ubiquitin-protein ligase Nedd4-2 (neural precursor cell expressed, developmentally downregulated 4-2). Overexpression of Nedd4-2 enhanced OAT3 ubiquitination, decreased OAT3 expression at the cell surface, and inhibited OAT3 transport activity. In contrast, overexpression of the ubiquitin ligase-dead mutant Nedd4-2/C821A or siRNA knockdown of endogenous Nedd4-2 had opposite effects on OAT3. Furthermore, immunoprecipitation experiments conducted both in culture cells and with rat kidney slices showed that there was a physical interaction between OAT3 and Nedd4-2. In conclusion, our results provided the first evidence that Nedd4-2 is an important regulator for OAT3 ubiquitination, expression, and transport activity.