Up-regulation of Na,K-ATPase β1 Transcription by Hyperoxia Is Mediated by SP1/SP3 Binding*

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 β1transcription 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.

[1]  H. Towle,et al.  Hyperoxia upregulated Na,K-adenosine triphosphatase beta1 gene transcription. , 1999, Chest.

[2]  R. Menon,et al.  Sp1 and Sp3 Regulate Transcriptional Activity of the Facilitative Glucose Transporter Isoform-3 Gene in Mammalian Neuroblasts and Trophoblasts* , 1998, The Journal of Biological Chemistry.

[3]  K. Webster,et al.  Hypoxia Regulates β-Enolase and Pyruvate Kinase-M Promoters by Modulating Sp1/Sp3 Binding to a Conserved GC Element* , 1998, The Journal of Biological Chemistry.

[4]  P. Factor,et al.  Augmentation of lung liquid clearance via adenovirus-mediated transfer of a Na,K-ATPase beta1 subunit gene. , 1998, The Journal of clinical investigation.

[5]  L. Aiello,et al.  Transcription Factors Sp1 and Sp3 Alter Vascular Endothelial Growth Factor Receptor Expression through a Novel Recognition Sequence* , 1998, The Journal of Biological Chemistry.

[6]  M. Runge,et al.  Differential transcriptional regulation of the human thrombin receptor gene by the Sp family of transcription factors in human endothelial cells. , 1998, The Biochemical journal.

[7]  H. Towle,et al.  Regulation of Na-K-ATPase gene expression by hyperoxia in MDCK cells. , 1998, American journal of physiology. Cell physiology.

[8]  U. Ikeda,et al.  Promoter of the Na,K-ATPase alpha3 subunit gene is composed of cis elements to which NF-Y and Sp1/Sp3 bind in rat cardiocytes. , 1997, Biochimica et biophysica acta.

[9]  A. Bell,et al.  Transcriptional regulation of the hepatocyte growth factor (HGF) gene by the Sp family of transcription factors , 1997, Oncogene.

[10]  Frank Grosveld,et al.  Transcription Factor Sp1 Is Essential for Early Embryonic Development but Dispensable for Cell Growth and Differentiation , 1997, Cell.

[11]  A. P. Butler,et al.  Transcription factor Sp3 antagonizes activation of the ornithine decarboxylase promoter by Sp1. , 1997, Nucleic acids research.

[12]  C. Weigert,et al.  Role of the stimulatory proteins Sp1 and Sp3 in the regulation of transcription of the rat pyruvate kinase M gene. , 1997, European journal of biochemistry.

[13]  Doris Schäfer,et al.  Redox‐regulated expression of glycolytic enzymes in resting and proliferating rat thymocytes , 1997, FEBS letters.

[14]  D. Ingbar,et al.  Cell density and antioxidant vitamins determine the effects of hyperoxia on proliferation and death of MDCK epithelial cells. , 1997, Nutrition and cancer.

[15]  M. Beato,et al.  An inhibitor domain in Sp3 regulates its glutamine‐rich activation domains. , 1996, The EMBO journal.

[16]  R. Pinkus,et al.  Role of Oxidants and Antioxidants in the Induction of AP-1, NF-κB, and Glutathione S-Transferase Gene Expression* , 1996, The Journal of Biological Chemistry.

[17]  C. Bingle,et al.  The lung enriched transcription factor TTF-1 and the ubiquitously expressed proteins Sp1 and Sp3 interact with elements located in the minimal promoter of the rat Clara cell secretory protein gene. , 1996, The Biochemical journal.

[18]  L. Gottlieb,et al.  Hyperbaric oxygenation upregulates rat lung Na,K-ATPase. , 1996, The European respiratory journal.

[19]  K. Webster,et al.  Physical and functional sensitivity of zinc finger transcription factors to redox change , 1996, Molecular and cellular biology.

[20]  G. Stein,et al.  Sp1 trans-activation of cell cycle regulated promoters is selectively repressed by Sp3. , 1995, Biochemistry.

[21]  M. Beato,et al.  Functional Analyses of the Transcription Factor Sp4 Reveal Properties Distinct from Sp1 and Sp3 (*) , 1995, The Journal of Biological Chemistry.

[22]  M. Beato,et al.  Sp1‐mediated transcriptional activation is repressed by Sp3. , 1994, The EMBO journal.

[23]  H. Pahl,et al.  Regulation of the transcription factors NF-kappa B and AP-1 by redox changes. , 1994, Chemico-biological interactions.

[24]  G. Gick,et al.  Characterization of the 5' flanking region of the rat Na+/K(+)-ATPase beta 1 subunit gene. , 1992, Biochimica et biophysica acta.

[25]  D. Dreher,et al.  Hyperoxia induces alkalinization and dome formation in MDCK epithelial cells. , 1992, The American journal of physiology.

[26]  K. Geering.  The functional role of the β‐subunit in the maturation and intracellular transport of Na,K‐ATPase , 1991 .

[27]  T. Rushmore,et al.  The antioxidant responsive element. Activation by oxidative stress and identification of the DNA consensus sequence required for functional activity. , 1991, The Journal of biological chemistry.

[28]  B. Rossier,et al.  Structure-function relationship of Na,K-ATPase. , 1991, Annual review of physiology.

[29]  J. Lingrel,et al.  Molecular genetics of Na,K-ATPase. , 1990, Progress in nucleic acid research and molecular biology.

[30]  J. Skou,et al.  [1] Overview: The Na,K-pump , 1988 .