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.
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.
Mesh Terms:
Animals, Binding Sites, Blotting, Northern, Blotting, Western, Carrier Proteins, Cell Line, DNA, Complementary, DNA-Binding Proteins, Dexamethasone, Down-Regulation, Epithelial Cells, Epithelial Sodium Channel, HMGA2 Protein, High Mobility Group Proteins, Mitogen-Activated Protein Kinases, Models, Biological, Plasmids, Precipitin Tests, Protein Binding, Rats, Receptors, Glucocorticoid, STAT3 Transcription Factor, Salivary Glands, Signal Transduction, Sodium Channels, Trans-Activators, Transcription, Genetic, Transcriptional Activation, Transfection, Two-Hybrid System Techniques, ras Proteins
Animals, Binding Sites, Blotting, Northern, Blotting, Western, Carrier Proteins, Cell Line, DNA, Complementary, DNA-Binding Proteins, Dexamethasone, Down-Regulation, Epithelial Cells, Epithelial Sodium Channel, HMGA2 Protein, High Mobility Group Proteins, Mitogen-Activated Protein Kinases, Models, Biological, Plasmids, Precipitin Tests, Protein Binding, Rats, Receptors, Glucocorticoid, STAT3 Transcription Factor, Salivary Glands, Signal Transduction, Sodium Channels, Trans-Activators, Transcription, Genetic, Transcriptional Activation, Transfection, Two-Hybrid System Techniques, ras Proteins
J. Biol. Chem.
Date: Aug. 10, 2001
PubMed ID: 11390395
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