Upstream sequences confer atrial-specific expression on the human atrial natriuretic factor gene.
5'-Flanking sequences from the human atrial natriuretic factor (hANF) gene were subcloned into a reporter plasmid (pSVOCAT) and transfected into primary cultures of neonatal rat atrial cardiocytes. Hybrid hANFCAT genes containing either 2500 or 409 base pairs of 5'-flanking sequence DNA were expressed at similar levels. When sequences between -409 and -332 were deleted, reporter gene (CAT) activity decreased significantly. Expression of the hANFCAT constructs was specific for atrial cells, as no expression was detected in primary cultures of ventricular cardiocytes or nonmyocardial cells derived from the neonatal hearts. Correct transcription start sites for the transfected hANF genes were confirmed by S1 nuclease mapping and RNase protection analysis. A "gel shift" assay was used to identify a specific cardiac nuclear protein which bound to the 5'-flanking sequence of the hANF gene. A 192-base pair PvuII fragment (-400 to -208) associated with a protein in these extracts in a tissue- and sequence-specific fashion. These findings indicate that the DNA sequence between -409 and -332 in the hANF gene harbors a tissue-specific element whose activity may involve association with a cardiac-specific nuclear protein.
LaPointe
M C
MC
Department of Medicine, University of California, San Francisco 94143.
Wu
J P
JP
Greenberg
B
B
Gardner
D G
DG
D000818:Animals; D000831:Animals, Newborn; D009320:Atrial Natriuretic Factor / Q000235:genetics*; D002478:Cells, Cultured; D003001:Cloning, Molecular; D005796:Genes; D005809:Genes, Regulator; D006325:Heart Atria / Q000378:metabolism; D006801:Humans; D009206:Myocardium / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism; D010957:Plasmids; D051381:Rats; D014158:Transcription, Genetic; D014162:Transfection
C472955:NPPA protein, human; D009687:Nuclear Proteins; D009320:Atrial Natriuretic Factor