@article {Zhangmol.112.082065, author = {Qiang Zhang and Shanshan Li and Cam Patterson and Guofeng You}, title = {Lysine 48-linked Polyubiquitination of Organic Anion Transporter-1 is Essential for Its Protein Kinase C-Regulated Endocytosis}, elocation-id = {mol.112.082065}, year = {2012}, doi = {10.1124/mol.112.082065}, publisher = {American Society for Pharmacology and Experimental Therapeutics}, abstract = {Organic anion transporter-1 (OAT1) mediates the body disposition of a diverse array of environmental toxins, and clinically important drugs. Therefore, understanding the regulation of this transporter has profound clinical significance. We previously established (Zhang Q, Hong M, Duan P, Pan Z, Ma J, You G (2008), J. Biol. Chem. 283:32570-9) that OAT1 undergoes constitutive internalization from and recycling back to cell surface and that acute activation of protein kinase C (PKC) inhibits OAT1 activity by reducing OAT1 cell surface expression through accelerating its internalization from cell surface to intracellular compartments. However, the underlying mechanisms are poorly understood. In the current study, we provided novel evidence that acute activation of PKC significantly enhanced OAT1 ubiquitination both in vitro and ex vivo. We further showed that ubiquitination of cell surface OAT1 increased in cells transfected with dominant negative mutant of dynamin-2, a maneuver blocking OAT1 internalization, suggesting that OAT1 ubiquitination proceeds before OAT1 internalization. Mass spectroscopy revealed that ubiquitination of OAT1 consisted of polyubiquitin chains primarily through lysine 48 (K48) linkage. Transfection of cells with dominant negative mutant of ubiquitin Ub-K48R, which prevents the formation of K48-linked polyubiquitin chains, abolished PKC-stimulated OAT1 ubiquitination and internalization. Together, our findings demonstrated for the first time that K48-linked polyubiquitination is essential for PKC-regulated OAT1 trafficking.}, issn = {0026-895X}, URL = {https://molpharm.aspetjournals.org/content/early/2012/10/19/mol.112.082065}, eprint = {https://molpharm.aspetjournals.org/content/early/2012/10/19/mol.112.082065.full.pdf}, journal = {Molecular Pharmacology} }