THE STRUCTURE OF THE PROSTAGLANDIN EP4 RECEPTOR GENE AND RELATED PSEUDOGENES

The EP4 prostaglandin receptor (EP4R) is a member of the seven transme mbrane receptor superfamily. We have obtained the human EP4 receptor g ene sequence and determined its structure relative to EP4R cDNA synthe sized from peripheral blood lymphocytes. The EP4R gene spans approxima tely 22 kb and consists of three exons separated by two introns. The f irst exon (530 bp) is noncoding. After an intron of 472 bp, the second exon contains a short (43 bp) 5' sequence before a 289-amino-acid ope n reading frame (ORF). An 11.5-kb intron is found at the end of transm embrane 6, and the rest of the ORF is in exon 3. The gene structure is analogous to those of the thromboxane, PGI, and PGD receptors. The de duced initiation site does not contain a conventional TATA box but is 70% GC-rich and contains CCAAT boxes, SP1 and AP2 motifs, and motifs c onsistent with activation by proinflammatory cytokines. Southern blot analysis of human genomic DNA shows two genes with homology to the EP4 R gene. Both appear to be pseudogenes with 70% amino acid identity to the EP4R up to the ''ERY'' sequence at the end of transmembrane 3, whe re an Alu-like repetitive sequence element was found. The ORF sequence is also interrupted by a stop codon. The pseudogenes differ in that o ne contains a second ''repetitive element'' (a line 1 repeat) in the 5 ' end of the ORF. Northern blot analysis of human mRNA using a pseudog ene probe showed hybridization only to the EP4 receptor transcript. PC R also failed to detect expression of either pseudogene. This study de fines the gene structure of EP4R and suggests the existence of two rel ated pseudogenes. (C) 1996 Academic Press, Inc.