Nuclear Factor kappa B Mediates Suppression of Canonical Transient Receptor Potential 6 Expression by Reactive Oxygen Species and Protein Kinase C in Kidney Cells

作者：Wang Yanxia; Ding Min; Chaudhari Sarika; Ding Yanfeng; Yuan Joseph; Stankowska Dorota; He Shaoqing; Krishnamoorthy Raghu; Cunningham Joseph T; Ma Rong^*

来源：JOURNAL OF BIOLOGICAL CHEMISTRY, 2013, 288(18): 12852-12865.

DOI：10.1074/jbc.M112.410357

This study was carried out to explore the molecular mechanism for down-regulation of TRPC6 expression in the reactive oxygen species (ROS)/PKC signaling in kidney cells. In cultured human mesangial cells, H2O2 and TNF-alpha inhibited TRPC6 mRNA expression in a time-dependent manner. Inhibition of NF-kappa B reversed both H2O2-and phorbol 12-myristate 13-acetate (PMA)-induced decrease in TRPC6 protein expression. Activation of NF-kappa B by knocking down I kappa B alpha using siRNA could mimic the suppressive effect of ROS/PKC on TRPC6. a Ca2+ imaging study showed that activation and inhibition of NF-kappa B significantly decreased and increased the TRPC6-mediated Ca2+ entry, respectively. Further experiments showed that PMA, but not its inactive analog 4 alpha-phorbol 12, 13-didecanoate (4 alpha-PDD), caused phosphorylation of I kappa B alpha and stimulated the nuclear translocation of NF-kappa B p50 and p65 subunits. The PMA-dependent I kappa B alpha phosphorylation was significantly inhibited by Go6976. Electrophoretic mobility shift assay revealed that PMA stimulated DNA binding activity of NF-kappa B. Furthermore, specific knockdown of p65, but not p50, prevented an H2O2 inhibitory effect on TRPC6 protein expression, suggesting p65 as a predominant NF-kappa B subunit repressing TRPC6. In agreement with a major role of p65, chromatin immunoprecipitation assays showed that PMA treatment induced p65 binding to the TRPC6 promoter. Moreover, PMA treatment increased the association of p65 with histone deacetylase (HDAC) and decreased histone acetylation at the TRPC6 promoter. Consistently, knockdown of HDAC2 by siRNA or inhibition of HDAC with trichostatin A prevented a H2O2-induced decrease in TRPC6 mRNA and protein expressions, respectively. Taken together, our findings imply an important role of NF-kappa B in a negative regulation of TRPC6 expression at the gene transcription level in kidney cells.

• 出版日期2013-5-3