Structure of the distal human gonadotropin releasing hormone (hGnRH) gene promoter and functional analysis in GT1-7 neuronal cells

Jadwiga K. Kepa, Andrea J. Spaulding, Britta M. Jacobsen, Zhaoqin Fang, Xiaoyan Xiong, Sally Radovick, Margaret E. Wierman

Research output: Contribution to journalArticle

Abstract

To assess potential species-specific expression of gonadotropin releasing hormone (GnRH), the distal human (h) GnRH promoter was cloned, characterized and tested in gene transfer studies. The nucleotide sequence of ~ 3.8 kb of 5'-flanking region was determined. Homology to the rat (r) GnRH sequence was observed in the proximal promoter region between -551 h (-424 r) and the transcriptional start site and within multiple distal promoter regions. In contrast, there was little similarity in the sequences between -1131/-551 h and -1031/-424 r. A deletion panel of 5'-flanking hGnRH promoter constructs was made and tested in transient transfection assays in GnRH-producing mouse GT1-7 neuronal cells. The largest hGnRH promoter construct (-3832/+5 h) exhibited high levels of reporter activity, similar to that observed with the largest rGnRH construct (-3026/+116 r). However, in contrast to the rat gene, deletion of distal promoter sequences of the hGnRH promoter to -1971, -1131 or -551 did not result in a decrease in luciferase reporter activity. Further truncation to -350 resulted in a 3-fold decrease in luciferase activity. There was no preferential use of the putative upstream hGnRH start site in neuronal cells. DNase I protection assays showed unique protection patterns with nuclear extracts from GT1-7 and Gn10 neuronal cells and the hGnRH and rGnRH promoter fragments. These data suggest the presence of different cis-acting elements and transacting factors that mediate species-specific neuronal GnRH expression.

Original languageEnglish (US)
Pages (from-to)3614-3620
Number of pages7
JournalNucleic Acids Research
Volume24
Issue number18
StatePublished - 1996
Externally publishedYes

ASJC Scopus subject areas

  • Genetics

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    Kepa, J. K., Spaulding, A. J., Jacobsen, B. M., Fang, Z., Xiong, X., Radovick, S., & Wierman, M. E. (1996). Structure of the distal human gonadotropin releasing hormone (hGnRH) gene promoter and functional analysis in GT1-7 neuronal cells. Nucleic Acids Research, 24(18), 3614-3620.