The transcription factor CREMtau and cAMP regulate promoter activity of the Na,K-ATPase alpha4 isoform.
The Na,K-ATPase is an essential enzyme of the plasma membrane that plays a key role in numerous cell processes that depend on the transcellular gradients of Na(+) and K(+). Among the various isoforms of the catalytic subunit of the Na,K-ATPase, alpha4 exhibits the most limited pattern of expression, being restricted to male germ cells. Activity of alpha4 is essential for sperm function, and alpha4 is upregulated during spermatogenesis. The present study addressed the transcriptional control of the human Na,K-ATPase alpha4 gene, ATP1A4. We describe that a 5' untranslated region of the ATP1A4 gene (designated -339/+480 based on the ATP1A4 transcription initiation site) has promoter activity in luciferase reporter assays. Computer analysis of this promoter region revealed consensus sites (CRE) for the cyclic AMP (cAMP) response element modulator (CREM). Accordingly, dibutyryl cAMP (db-cAMP) and ectopic expression of CREMtau, a testis specific splice variant of CREM were able to activate the ATP1A4 promoter driven expression of luciferase in HEK 293 T, JEG-3 and GC-1 cells. Further characterization of the effect of db-cAMP and CREMtau on deleted constructs of the ATP1A4 promoter (-339/+80, and +25/+480), and on the -339/+480 region carrying mutations in the CRE sites showed that db-cAMP and CREMtau effect required the CRE motif located 263 bp upstream the transcription initiation site. EMSA experiments confirmed the CRE sequence as a bonafide CREMtau binding site. These results constitute the first demonstration of the transcriptional control of ATP1A4 gene expression by cAMP and by CREMtau, a transcription factor essential for male germ cell gene expression.
Rodova
Marianna
M
Department of Molecular and Integrative Physiology, University of Kansas Medical Center, Kansas City, Kansas 66160, USA.
Nguyen
Anh-Nguyet
AN
Blanco
Gustavo
G
D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism*; D051786:Cyclic AMP Response Element Modulator / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D018507:Gene Expression Regulation, Developmental; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D009154:Mutation; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D000254:Sodium-Potassium-Exchanging ATPase / Q000235:genetics*; D013091:Spermatogenesis / Q000235:genetics*; D014157:Transcription Factors / Q000378:metabolism*; D024363:Transcription Initiation Site
D012333:RNA, Messenger; D014157:Transcription Factors; D051786:Cyclic AMP Response Element Modulator; D000242:Cyclic AMP; C485719:ATP1A4 protein, human; D000254:Sodium-Potassium-Exchanging ATPase