Isolation of a bi-directional promoter directing expression of the mouse GABP alpha and ATP synthase coupling factor 6 genes

The GA-binding protein (GABP) is a ubiquitous heteromeric transcription fac tor implicated in the regulation of several genes involved in mitochondrial energy metabolism including subunits of cytochrome c oxidase, ATP synthase , and mitochondrial transcription factor 1 (mtTF1). GABP alpha subunit bind s the PEA3/Ets binding sites (EBS), while GABP beta contains a transcriptio n activation domain and mediates alpha beta dimer and alpha (2)beta (2) tet ramer formation essential for activation of transcription. Here we report t he cloning of 2449 bp of the mouse (m) GABP alpha promoter region including 201 bp of the 5' end of the published mGABP alpha cDNA sequence. Surprisin gly, sequences homologous to the 5'UTR of mouse, rat and human mitochondria l ATP synthase coupling factor 6 (ATPsynCF6) cDNAs were found165-240 bp ups tream of the mGABP alpha cDNA. A search of the non-redundant nucleotide dat abase revealed a human genomic sequence derived from chromosome 21 (21q22) bearing significant homology to the mGABP alpha /ATPsynCF6 promoter region and encompassed the entire hGABP alpha and hATP-synCF6 genes. Primer extens ion analysis revealed multiple transcription start sites for both mGABP alp ha and mATPsynCF6 mRNAs that mapped near the published cDNA 5' ends. Sequen ce analysis identified several binding sites upstream of the GABP alpha cDN A sequence including sites for GABP (-86, -104, -169, -257, and -994), YY1 (-57), Spl (-242 and -226), and NRF1 (-5). No 'TATA' motif was identified n ear either the GABPa or ATPsynCF6 transcription start sites. The human and mouse promoters retain significant sequence identity including binding site s for several tissue-specific transcription factors. Transient transfection assays using Luciferase reporter constructs containing the intergenic regi on and flanking sequences confirmed that this region of DNA promotes transc ription in both directions. (C) 2000 Elsevier Science B.V. All rights reser ved.