title: Rottlerin inhibits tonicity-dependent expression and action of TonEBP in a PKCdelta-independent fashion.
abstract: Novel protein kinase C (PKC) isoforms PKCdelta and PKCepsilon have recently been implicated in signaling by hypertonic stress. We investigated the role of the putative PKCdelta inhibitor rottlerin on tonicity-dependent gene regulation. In the renal medullary mIMCD3 cell line, rottlerin blocked tonicity-dependent transcription of a tonicity enhancer (TonE)-driven luciferase reporter gene, as well as tonicity-dependent transcription of the physiological tonicity effector gene aldose reductase, but not urea-dependent transcription. Consistent with these data, rottlerin inhibited tonicity-dependent expression of TonE binding protein (TonEBP) at the mRNA and protein levels. Another inhibitor of both novel and conventional PKC isoforms, GF-109203X, suppressed TonEBP-dependent transcription but failed to influence tonicity-inducible TonEBP expression. Global PKC downregulation with protracted phorbol ester treatment, however, failed to influence tonicity-dependent signaling, arguing against a PKCdelta-dependent mechanism of rottlerin action in this model. In addition, hypertonic stress failed to induce phosphorylation of PKCdelta. Furthermore, in a PC-12 cell model with a comparable degree of tonicity-dependent transcription, constitutive overexpression of dominant negative-acting PKCdelta or PKCepsilon effectively decreased tonicity signaling to extracellular signal-regulated kinase activation, as expected, but failed to influence TonE-dependent transcription. TonE-dependent transcription, however, remained rottlerin sensitive in this PC-12 cell model. In the aggregate, these data indicate that rottlerin dramatically inhibits tonicity-dependent TonEBP expression and TonE-dependent transcription but, despite its reputed mode of action, does so through a PKCdelta-independent pathway.
authors.affiliation: Division of Nephrology and Department of Cell and Developmental Biology, Oregon Health and Science University and the Portland Veterans Affairs Medical Center, Portland, Oregon 97201, USA.
authors.forename: Hongyu
authors.initials: H
authors.lastname: Zhao
authors.forename: Wei
authors.initials: W
authors.lastname: Tian
authors.forename: David M
authors.initials: DM
authors.lastname: Cohen
mesh_terms: D000098:Acetophenones / Q000494:pharmacology*; D000449:Aldehyde Reductase / Q000096:biosynthesis / Q000235:genetics; D000818:Animals; D001578:Benzopyrans / Q000494:pharmacology*; D015153:Blotting, Western; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005347:Fibroblasts / Q000187:drug effects / Q000502:physiology; D017930:Genes, Reporter; D007093:Imidazoles / Q000494:pharmacology; D007527:Isoenzymes / Q000037:antagonists & inhibitors*; D008156:Luciferases / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors*; D051745:Protein Kinase C-delta; D011725:Pyridines / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015534:Trans-Activators / Q000037:antagonists & inhibitors* / Q000096:biosynthesis* / Q000235:genetics; D014157:Transcription Factors; D014158:Transcription, Genetic / Q000187:drug effects
chemical_list: D000098:Acetophenones; D001578:Benzopyrans; D004791:Enzyme Inhibitors; D007093:Imidazoles; D007527:Isoenzymes; C492200:Nfat5 protein, mouse; D011725:Pyridines; D012333:RNA, Messenger; D015534:Trans-Activators; D014157:Transcription Factors; C085746:rottlerin; D000449:Aldehyde Reductase; D008156:Luciferases; C497259:Prkcd protein, mouse; D011493:Protein Kinase C; D051745:Protein Kinase C-delta; C093642:SB 203580