Transcriptional Activation of the Human Hematopoietic Prostaglandin D Synthase Gene in Megakaryoblastic Cells: ROLES OF THE Oct-1 ELEMENT IN THE 5′-FLANKING REGION AND THE AP-2 ELEMENT IN THE UNTRANSLATED EXON 1

Ko Fujimori,Yoshihide Kanaoka,Yuriko Sakaguchi,Yoshihiro Urade

doi:10.1074/jbc.m007688200

Ko Fujimori, Yoshihide Kanaoka + Show 2 more

Open Access

https://doi.org/10.1074/jbc.m007688200

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Abstract

The human hematopoietic prostaglandin D synthase (H-PGDS) gene is highly expressed in human megakaryoblastic cells, in which phorbol ester induces its expression. We characterized the promoter activity of the 5'-flanking region and the untranslated exon 1 (-1044 to +290) of the human H-PGDS gene in human megakaryoblastic Dami cells. Transient expression analysis using the luciferase reporter gene revealed that the 5'-flanking region and the untranslated exon 1 were sufficient for efficient expression of the H-PGDS gene in Dami cells, but not in monocytic U937 cells. Deletion and site-directed mutagenesis of the Oct-1 element in the 5'-flanking region decreased the promoter activity by approximately 30% compared with that of the entire region from -1044 to +290. An electrophoretic mobility shift assay demonstrated that Oct-1 specifically bound to the promoter region. Interestingly, even only untranslated exon 1 (+1 to +290) showed approximately 60% of the promoter activity of the entire region from -1044 to +290. Site-directed mutagenesis of the AP-2 element within the untranslated exon 1 abolished the basal promoter activity as well as its phorbol ester-mediated up-regulation. In AP-2-deficient HepG2 cells, the H-PGDS promoter activity was enhanced by coexpression with AP-2alpha. These findings indicate that the Oct-1 element in the 5'-flanking region acts as a positive cis-acting element and that the AP-2 element in the untranslated exon 1 is crucial for both basal and phorbol ester-mediated up-regulation of human H-PGDS gene expression in megakaryoblastic Dami cells.

Highlights

The human hematopoietic prostaglandin D synthase (H-PGDS) gene is highly expressed in human megakaryoblastic cells, in which phorbol ester induces its expression
In AP2-deficient HepG2 cells, the H-PGDS promoter activity was enhanced by coexpression with AP-2␣. These findings indicate that the Oct-1 element in the 5؅-flanking region acts as a positive cis-acting element and that the AP-2 element in the untranslated exon 1 is crucial for both basal and phorbol ester-mediated up-regulation of human H-PGDS gene expression in megakaryoblastic Dami cells
The 5Ј-flanking region and the untranslated exon 1 of the human H-PGDS gene contained several potential cis-acting elements recognized by the transcription factors that may play roles in the regulation of the human H-PGDS gene, including one Oct-1 element, five GATA elements

Summary

Introduction

The human hematopoietic prostaglandin D synthase (H-PGDS) gene is highly expressed in human megakaryoblastic cells, in which phorbol ester induces its expression. These findings indicate that the Oct-1 element in the 5؅-flanking region acts as a positive cis-acting element and that the AP-2 element in the untranslated exon 1 is crucial for both basal and phorbol ester-mediated up-regulation of human H-PGDS gene expression in megakaryoblastic Dami cells.

Results

Conclusion