title: Nuclear factor κB mediates suppression of canonical transient receptor potential 6 expression by reactive oxygen species and protein kinase C in kidney cells.
abstract: 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-α inhibited TRPC6 mRNA expression in a time-dependent manner. Inhibition of NF-κB reversed both H2O2- and phorbol 12-myristate 13-acetate (PMA)-induced decrease in TRPC6 protein expression. Activation of NF-κB by knocking down IκBα using siRNA could mimic the suppressive effect of ROS/PKC on TRPC6. a Ca(2+) imaging study showed that activation and inhibition of NF-κB significantly decreased and increased the TRPC6-mediated Ca(2+) entry, respectively. Further experiments showed that PMA, but not its inactive analog 4α-phorbol 12, 13-didecanoate (4α-PDD), caused phosphorylation of IκBα and stimulated the nuclear translocation of NF-κB p50 and p65 subunits. The PMA-dependent IκBα phosphorylation was significantly inhibited by Gö6976. Electrophoretic mobility shift assay revealed that PMA stimulated DNA binding activity of NF-κB. Furthermore, specific knockdown of p65, but not p50, prevented an H2O2 inhibitory effect on TRPC6 protein expression, suggesting p65 as a predominant NF-κB 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-κB in a negative regulation of TRPC6 expression at the gene transcription level in kidney cells.
authors.affiliation: Department of Integrative Physiology and Cardiovascular Research Institute, University of North Texas Health Science Center, Fort Worth, Texas 76107, USA.
authors.forename: Yanxia
authors.initials: Y
authors.lastname: Wang
authors.forename: Min
authors.initials: M
authors.lastname: Ding
authors.forename: Sarika
authors.initials: S
authors.lastname: Chaudhari
authors.forename: Yanfeng
authors.initials: Y
authors.lastname: Ding
authors.forename: Joseph
authors.initials: J
authors.lastname: Yuan
authors.forename: Dorota
authors.initials: D
authors.lastname: Stankowska
authors.forename: Shaoqing
authors.initials: S
authors.lastname: He
authors.forename: Raghu
authors.initials: R
authors.lastname: Krishnamoorthy
authors.forename: Joseph T
authors.initials: JT
authors.lastname: Cunningham
authors.forename: Rong
authors.initials: R
authors.lastname: Ma
mesh_terms: D002227:Carbazoles / Q000494:pharmacology; D002273:Carcinogens / Q000494:pharmacology; D002460:Cell Line; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects* / Q000502:physiology; D055785:Gene Knockdown Techniques; D056464:Histone Deacetylase 2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D056572:Histone Deacetylase Inhibitors / Q000494:pharmacology; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D006877:Hydroxamic Acids / Q000494:pharmacology; D007668:Kidney / Q000166:cytology / Q000378:metabolism*; D052002:NF-kappa B p50 Subunit / Q000235:genetics / Q000378:metabolism*; D016877:Oxidants / Q000494:pharmacology*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D020218:Response Elements / Q000502:physiology; D050052:TRPC Cation Channels / Q000096:biosynthesis* / Q000235:genetics; D000074287:TRPC6 Cation Channel; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D051996:Transcription Factor RelA / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology
chemical_list: D002227:Carbazoles; D002273:Carcinogens; D004791:Enzyme Inhibitors; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; D052002:NF-kappa B p50 Subunit; C489548:NFKB1 protein, human; D016877:Oxidants; C498143:RELA protein, human; D050052:TRPC Cation Channels; D000074287:TRPC6 Cation Channel; C117032:TRPC6 protein, human; D051996:Transcription Factor RelA; C081021:Go 6976; C012589:trichostatin A; D006861:Hydrogen Peroxide; D011493:Protein Kinase C; C533423:HDAC2 protein, human; D056464:Histone Deacetylase 2; D013755:Tetradecanoylphorbol Acetate
keywords: Calcium; Hydrogen Peroxide; Ion Channels; NF-κB; Reactive Oxygen Species (ROS); TRP Channels