Cloning of the mouse Pax4 gene promoter and identification of a pancreatic beta cell specific enhancer.
Pax4 encodes a paired-box transcription factor and is essential for the differentiation of islet cells since the Pax4 homozygous mutant mice lack mature beta and delta cells. However, little is known about the transcriptional regulation of the Pax4 gene. We isolated and sequenced a 2.4-kb mouse genomic DNA fragment containing the 5' flanking sequence of Pax4 and identified a previously unrecognized intron. Primer extension revealed that this TATA-less promoter had only one transcription start site. The promoter activity of this fragment with various deletion mutants when tested in beta and non-beta cell lines indicated the presence of a beta-cell specific enhancer in the region, -1858 to -1954 bp. DNase 1 footprinting and gel retardation assays indicated that nuclear proteins from betaHC3 cells interacted with two sequences which contained putative CdxA/Nkx.2 and GATA-1,-2 binding sites. Site-directed mutagenesis indicated that both of these regions were necessary for beta-cell specific enhancer activity.
Xu
W
W
Department of Physiology, University of Manitoba, Winnipeg, Canada R3E 0W3.
Murphy
L J
LJ
D020121:5' Untranslated Regions / Q000032:analysis / Q000235:genetics; D000818:Animals; D003001:Cloning, Molecular; D018387:Codon, Initiator; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D018398:Homeodomain Proteins / Q000235:genetics*; D006801:Humans; D007515:Islets of Langerhans / Q000378:metabolism*; D051379:Mice; D016297:Mutagenesis, Site-Directed; D051761:Paired Box Transcription Factors; D011401:Promoter Regions, Genetic / Q000235:genetics*; D051381:Rats; D017384:Sequence Deletion; D014157:Transcription Factors / Q000235:genetics*; D014162:Transfection; D014407:Tumor Cells, Cultured
D020121:5' Untranslated Regions; D018387:Codon, Initiator; D018398:Homeodomain Proteins; C491709:PAX4 protein, human; D051761:Paired Box Transcription Factors; C110218:Pax4 protein, mouse; C491708:Pax4 protein, rat; D014157:Transcription Factors