Up-regulation of Na,K-ATPase β1 transcription by hyperoxia is mediated by SP1/SP3 binding

Christine H. Wendt, Greg Gick, Renuka Sharma, Yong Zhuang, Wenlian Deng, David H. Ingbar

Research output: Contribution to journalArticlepeer-review

24 Scopus citations

Abstract

The sodium pump, Na,K-ATPase, is an important protein for maintaining intracellular ion concentration, cellular volume, and ion transport and is regulated both transcriptionally and post-transcriptionally. We previously demonstrated that hyperoxia increased Na,K-ATPase β1 gene expression in Madin-Darby canine kidney (MDCK) cells. In this study, we identify a DNA element necessary for up-regulation of the Na,K-ATPase β1 transcription by hyperoxia and evaluate the nuclear proteins responsible for this up-regulation. Transient transfection experiments in MDCK cells using sequential 5′-deletions of the rat Na,K-ATPase β1 promoter-luciferase fusion gene demonstrated promoter activation by hyperoxia between -102 and +151. The hyperoxia response was localized to a 7-base pair region between -62 and -55, which contained a GC-rich region consistent with a consensus sequence for the SP1 family, that was sufficient for up-regulation by hyperoxia. This GC element exhibited both basal and hyperoxia-induced promoter activity and bound both transcription factors SP1 and SP3 in electrophoretic mobility shift assays. In addition, electrophoretic mobility shift assays demonstrated increased binding of SP1/SP3 in cells exposed to hyperoxia while mutation of this element eliminated protein binding. Other GC sites within the proximal promoter also demonstrated up-regulation of transcription by hyperoxia, however, the site at -55 had higher affinity for SP proteins.

Original languageEnglish (US)
Pages (from-to)41396-41404
Number of pages9
JournalJournal of Biological Chemistry
Volume275
Issue number52
DOIs
StatePublished - Dec 29 2000

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