title: Requirement for high mobility group protein HMGI-C interaction with STAT3 inhibitor PIAS3 in repression of alpha-subunit of epithelial Na+ channel (alpha-ENaC) transcription by Ras activation in salivary epithelial cells.
abstract: Previously, we have demonstrated that oxidative stress or Ras/ERK activation leads to the transcriptional repression of alpha-subunit of epithelial Na(+) channel (ENaC) in lung and salivary epithelial cells. Here, we further investigated the coordinated molecular mechanisms by which alpha-ENaC expression is regulated. Using both stable and transient transfection assays, we demonstrate that the overexpression of high mobility group protein I-C (HMGI-C), a Ras/ERK-inducible HMG-I family member, represses glucocorticoid receptor (GR)/dexamethasone (Dex)-stimulated alpha-ENaC/reporter activity in salivary epithelial cells. Northern analyses further confirm that the expression of endogenous alpha-ENaC gene in salivary Pa-4 cells is suppressed by an ectopic HMGI-C overexpression. Through yeast two-hybrid screening and co-immunoprecipitation assays from eukaryotic cells, we also discovered the interaction between HMGI-C and PIAS3 (protein inhibitor of activated STAT3 (signal transducer and activator of transcription 3)). A low level of ectopically expressed PIAS3 cooperatively inhibits GR/Dex-dependent alpha-ENaC transcription in the presence of HMGI-C. Reciprocally, HMGI-C expression also coordinately enhances PIAS3-mediated repression of STAT3-dependent transactivation. Moreover, overexpression of antisense HMGI-C construct is capable of reversing the repression mediated by Ras V12 on GR- and STAT3-dependent transcriptional activation. Together, our results demonstrate that Ras/ERK-mediated induction of HMGI-C is required to effectively repress GR/Dex-stimulated transcription of alpha-ENaC gene and STAT3-mediated transactivation. These findings delineate a network of inhibitory signaling pathways that converge on HMGI-C.PIAS3 complex, causally associating Ras/ERK activation with the repression of both GR and STAT3 signaling pathways in salivary epithelial cells.
authors.affiliation: Department of Molecular Pharmacology and Toxicology, Will Rogers Institute, Pulmonary Research Center, University of Southern California, Los Angeles, California 90033, USA.
authors.forename: M D
authors.initials: MD
authors.lastname: Zentner
authors.forename: H H
authors.initials: HH
authors.lastname: Lin
authors.forename: H T
authors.initials: HT
authors.lastname: Deng
authors.forename: K J
authors.initials: KJ
authors.lastname: Kim
authors.forename: H M
authors.initials: HM
authors.lastname: Shih
authors.forename: D K
authors.initials: DK
authors.lastname: Ann
mesh_terms: D000818:Animals; D001665:Binding Sites; D015152:Blotting, Northern; D015153:Blotting, Western; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D018076:DNA, Complementary / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors*; D003907:Dexamethasone / Q000494:pharmacology; D015536:Down-Regulation; D004847:Epithelial Cells / Q000378:metabolism*; D053503:Epithelial Sodium Channels; D025743:HMGA2 Protein; D006609:High Mobility Group Proteins / Q000378:metabolism* / Q000502:physiology*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D008954:Models, Biological; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D051381:Rats; D011965:Receptors, Glucocorticoid / Q000096:biosynthesis; D050796:STAT3 Transcription Factor; D012469:Salivary Glands / Q000378:metabolism*; D015398:Signal Transduction; D015222:Sodium Channels / Q000378:metabolism*; D015534:Trans-Activators / Q000037:antagonists & inhibitors*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D020798:Two-Hybrid System Techniques; D018631:ras Proteins / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D053503:Epithelial Sodium Channels; D025743:HMGA2 Protein; D006609:High Mobility Group Proteins; D011965:Receptors, Glucocorticoid; D050796:STAT3 Transcription Factor; D015222:Sodium Channels; C494088:Stat3 protein, rat; D015534:Trans-Activators; D003907:Dexamethasone; D020928:Mitogen-Activated Protein Kinases; D018631:ras Proteins