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We performed cDNA and genomic cloning, sequencing and promoter analysis of the putative human O-sialoglycoprotein endopeptidase gene OSGEP (a homologue of gcp, a Pasteurella haemolytica A1 glycoprotease). The cloned OSGEP cDNA is 1311 nucleotides long, and encodes a protein consisting of 335 amino acids with predicted molecular mass of 36.4 kDa. The amino acid sequence of OSGEP showed 29.7% identity with that of P. haemolytica glycoprotease. The OSGEP gene is 7.75 kb long, consists of 11 exons and 10 introns, and lies immediately adjacent to the APEX gene (which encodes APEX nuclease, a multifunctional DNA repair enzyme) in 5′-to-5′ orientation. The promoter region of the OSGEP gene lacks the typical TATA box, but has putative regulatory elements in the CpG island. Northern blot analysis showed ubiquitous expression of the OSGEP gene in several tissues, and we observed similarities in expression patterns between OSGEP and APEX. In order to study the regulation of OSGEP gene expression, we analyzed the OSGEP promoter region by luciferase assay using HeLa cells. A functional region required for full transcription activity was narrowed down to a 23 bp region containing a CCAAT box. It has been reported that this CCAAT box promotes basal transcription in the APEX direction. We thus conclude that a bidirectional promoter containing a CCAAT box regulates transcription of both the OSGEP and APEX genes. [Copyright &y& Elsevier]

Abstract: Flap endonuclease 1 (FEN1) is a structure-specific nuclease involved in DNA replication and repair. The mouse Fen1 gene, which has two exons, is located immediately adjacent to the gene corresponding to full-length cDNA of Riken 1810006K21 (1810006K21Rik) in a head-to-head orientation. Transcription initiation sites of each gene are 274 bp apart in the mouse genome. The spacer sequence between the bidirectional genes contains a CpG island, but lacks the typical TATA box. In the present study, transcription of the mFen1 gene was started from two initiation sites, and the first noncoding exon was spliced to the second exon using two different splicing donor sites, producing 3 kinds of mFen1 transcripts. A 594-bp fragment between the mFen1 and 1810006K21Rik genes, which contains two conserved sequence blocks (CSB) between mouse and human sequence, functions as a bidirectional promoter. The multiple cis-elements, including an Ets-binding site and E-box in the CSB, are involved in activation or repression of transcription in both directions. Interestingly, the E-box activates mFen1 transcription and simultaneously represses promoter activity in the opposite direction. Mutation of either splicing donor site of the mFen1 gene produced limiting alternative splicing products, but did not affect luciferase activity. In contrast, the splicing-defective mutation produced by disruption of the acceptor site completely lacked luciferase activity, indicating that the splicing has a significant effect on production of luciferase protein by shortening the 5′-untranslated region of the mRNA. [Copyright &y& Elsevier]