AVP-induced VIT32 gene expression in collecting duct cells occurs via trans-activation of a CRE in the 5'-flanking region of the VIT32 gene

Christie P Thomas; Randy W Loftus; Kang Z Liu

doi:10.1152/ajprenal.00107.2004

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AVP-induced VIT32 gene expression in collecting duct cells occurs via trans-activation of a CRE in the 5'-flanking region of the VIT32 gene

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AVP-induced VIT32 gene expression in collecting duct cells occurs via trans-activation of a CRE in the 5'-flanking region of the VIT32 gene

Christie P Thomas, Randy W Loftus and Kang Z Liu

American journal of physiology. Renal physiology, Vol.287(3), pp.F460-468

09/2004

DOI: 10.1152/ajprenal.00107.2004

PMID: 15140762

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Abstract

VIT32, a vasopressin-induced transcript, inhibits Na(+) transport when coexpressed with the epithelial sodium channel in Xenopus laevis oocytes (EMBO J 21: 5109-5117, 2002). To understand the mechanism of VIT32 gene regulation, we examined the effect of DDAVP and cAMP stimulation on VIT32 expression in M-1 mouse collecting duct cells and in H441 human airway epithelial cells. Elevation of cAMP with forskolin and IBMX increased VIT32 gene expression with a peak effect at 2 h. The increase in gene expression was abolished by H89 and by actinomycin D, suggesting that cAMP stimulates VIT32 mRNA expression by a PKA-mediated increase in gene transcription. An approximately 1.5-kb fragment of the 5'-flanking region of VIT32 was cloned and was able to confer cAMP-stimulated reporter gene activity when transfected into M-1 and H441 cells. By deletion analysis and site-directed mutagenesis, a cAMP response element (CRE) was identified within the proximal promoter region that was sufficient to account for the increase in VIT32 gene expression seen with DDAVP and elevation of cAMP. Furthermore, DDAVP-stimulated VIT32 promoter-reporter activity was inhibited by H89 and by a dominant negative CREB construct. Finally, we were able to identify CREB as a nuclear protein that bound to the VIT32 CRE in gel mobility shift assays. In summary, DDAVP stimulates transcription of VIT32 via a CRE within the proximal promoter region of the VIT32 gene.

Cell Line

Gene Expression

Kidney Tubules, Collecting - physiology

Respiratory Mucosa - cytology

Humans

Mice, Inbred C57BL

Transcriptional Activation - drug effects

Molecular Sequence Data

Renal Agents - pharmacology

Kidney Tubules, Collecting - cytology

Carrier Proteins - genetics

Animals

Carrier Proteins - metabolism

Promoter Regions, Genetic - physiology

Base Sequence

Cyclic AMP Response Element-Binding Protein - metabolism

Mice

5' Flanking Region - physiology

Cyclic AMP - metabolism

Deamino Arginine Vasopressin - pharmacology

Details

Title: Subtitle: AVP-induced VIT32 gene expression in collecting duct cells occurs via trans-activation of a CRE in the 5'-flanking region of the VIT32 gene
Creators: Christie P Thomas - Department of Internal Medicine, Graduate Program in Molecular Biology, University of Iowa College of Medicine, Iowa City, Iowa, USA. christie-thomas@uiowa.edu
Randy W Loftus
Kang Z Liu
Resource Type: Journal article
Publication Details: American journal of physiology. Renal physiology, Vol.287(3), pp.F460-468
DOI: 10.1152/ajprenal.00107.2004
PMID: 15140762
NLM abbreviation: Am J Physiol Renal Physiol
ISSN: 1931-857X
eISSN: 1522-1466
Publisher: United States
Grant note: DK-54348 / NIDDK NIH HHS HL-71664 / NHLBI NIH HHS R01 DK054348 / NIDDK NIH HHS R01 HL071664 / NHLBI NIH HHS
Language: English
Date published: 09/2004
Academic Unit: Stead Family Department of Pediatrics; Anesthesia; Obstetrics and Gynecology; Internal Medicine
Record Identifier: 9983985903702771

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