Regulation of organic anion transporting polypeptide 1B1 transport function by lysine deacetylase 6.

Hepatic transport protein organic anion transporting polypeptide 1B1 (OATP1B1) is a key determinant of drug-drug interactions. We reported OATP1B1 lysine acetylation recently, however, the lysine deacetylase (KDAC), also known as histone deacetylase (HDAC), involved in its deacetylation remains uninvestigated. This study determined the role of KDAC6/HDAC6, a major cytosolic KDAC, on OATP1B1 acetylation and transport function. Loss-of-function of KDAC6 by CRISPR/Cas9-mediated knockout in HEK293T cells or by treatment with the selective KDAC6 inhibitor tubacin (TBC) (5 μM, 24 hours) in transporter-expressing HEK293 cells markedly reduces OATP1B1-mediated transport of [3H]estradiol-17-ß-D-glucuronide to 0.62 ± 0.095- and 0.28 ± 0.007-fold of control, respectively. TBC treatment did not affect OATP1B1 mRNA, protein levels and colocalization with plasma membrane marker Na/K-ATPase, suggesting a regulation at a post-translational level. TBC treatment also reduces [3H]rosuvastatin accumulation in primary human hepatocytes. Quantitative comparison of post-translational modifications (PTMs) between TBC treatment and control showed concurrent increase in lysine acetylation at K675 (to 5.16 ± 2.7-fold of control) and decrease in dual phosphorylation at Ser659-Ser663 (to 0.34 ± 0.13-fold of control). A variant S659A-S663A-K675Q-OATP1B1 that mimics these concurrent PTM changes reduces OATP1B1-mediated transport compared with the wild-type control, supporting a role for altered PTMs in downregulation of OATP1B1 transport function upon KDAC6 loss-of-function. In addition, K49, a residue important in maintaining OATP1B1 transport function, was identified as acetylated for the first time. This study identifies KDAC6 as a key enzyme regulating lysine acetylation of OATP1B1 and maintaining OATP1B1 transport function, thereby implying a novel KDAC6-linked mechanism in OATP1B1-mediated drug-drug interactions. SIGNIFICANCE STATEMENT: This study identifies lysine deacetylase 6 (KDAC6) as a key enzyme involved in post-translational regulation of OATP1B1. Current findings demonstrate that KDAC6-dependent acetylation-phosphorylation axis modulates OATP1B1 transport function and provides a mechanistic basis by which altered KDAC6 activity may influence OATP1B1-mediated drug-drug interactions.

Duke Scholars

Author Erik James Soderblom Cell Biology