Genomic structure and transcriptional regulation of the human NAD+-dependent 15-hydroxyprostaglandin dehydrogenase gene

Abstract

The NAD + -dependent 15-hydroxyprostaglandin dehydrogenase (PGDH) is a catabolic enzyme that controls the biological activities of prostaglandins by converting them into inactive keto-metabolites. Here we report the genomic organisation of the complete human PGDH gene and characterise its transcriptional regulation. The PGDH gene spans about 31 kb on chromosome 4 and contains 7 exons. Within 2·4 kb of the 5′-flanking sequence we identified two regions with clustered putative transcription factor binding sites. The distal promoter element PGDH-DE (positions−2152/−1944 relative to the start codon) contains binding sites for Ets and activating protein-1 (AP-1) flanked by two cAMP-responsive element-binding protein binding sites (CREB1, CREB2), whereas the proximal element PGDH-PE (−235/−153) includes an Ets and an AP-1 binding sequence. By electrophoretic mobility shift assay, no high affinity binding of Ets or AP-1 factors was observed with PGDH-PE, whereas we confirmed interaction of members of the Ets, AP-1 and CREB families of transcription factors with PGDH-DE. Transcriptional control of the PGDH promoter was assessed by transiently transfecting JEG-3 choriocarcinoma cells. A luciferase reporter gene construct containing the PGDH-PE was not induced by c-jun/c-fos in the absence or presence of co-expressed Ets-1. A construct carrying the PGDH-DE in front of the minimal homologous promoter was activated by co-transfection of expression vectors for AP-1 proteins. Mutation of the AP-1 or CREB2 site reduced the response to c-jun/c-fos, whereas mutation of the Ets site of the distal element reduced basal promoter activity. CREB activated the PGDH-DE construct through the CREB1 site. These results defined the distal element as an integrator of transcriptional regulation by AP-1, Ets and CREB proteins.

15-Hydroxyprostaglandin Dehydrogenase Is a Tumor Suppressor of Human Breast Cancer

“…Analysis of 10 primary breast cancers revealed methylation in three tumors. The methylation in primary tumors was less extensive than in MDA-MB-436 cells, but two areas were methylated in all three tumors, one near the TATA box and the other near a site of the methylation-sensitive transcription factor USF (13)(14)(15). Importantly, these tumors showed reduced expression of 15-PGDH mRNA compared with either MCF-7 cells (<25%) or the unmethylated tumors.…”

Levels of NAD+-dependent 15-hydroxyprostaglandin dehydrogenase are reduced in inflammatory bowel disease: evidence for involvement of TNF-α

“…Our studies suggest that TNF-␣ suppressed the transcription of 15-PGDH, resulting in reduced levels of 15-PGDH mRNA and protein and enzyme activity. Recently, AP-1, Ets, and CREB proteins have been implicated in the transcriptional regulation of 15-PGDH (26). Future studies will be needed to determine whether any one of these trans-activating factors is involved in TNF-␣-mediated suppression of 15-PGDH transcription.…”

Prognostic impact of transcription factor Fra-1 in ER-positive breast cancer: contribution to a metastatic phenotype through modulation of tumor cell adhesive properties

“…To generate stable clones with increased Fra-1 expression, the full Fra-1 cDNA that had been cloned before in pcDNA3.1+ plasmids was supplemented with a C-terminal FLAG epitope by PCR and subcloned in the bicistronic vector pIRES-P (Genbank no. Z75185) using the NheI and EcoR1 restriction enzymes (Nandy et al 2003). Transfection of MCF7 and MDA MB231 cells with this plasmid and the empty vector was performed with LipofectAMINE PLUS reagent (Life Technologies, Karlsruhe, Germany).…”