Nucleic Acids Research, Vol 24, Issue 18 3614-3620, Copyright © 1996 by Oxford University Press
JK Kepa, AJ Spaulding, BM Jacobsen, Z Fang, X Xiong, S Radovick and ME Wierman
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 approximately 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.
ARTICLES
Structure of the distal human gonadotropin releasing hormone (hGnrh) gene promoter and functional analysis in Gt1-7 neuronal cells
Department of Medicine, University of Colorado Health Sciences Center and Research Service, Veterans Affairs Medical Center, Denver 80220, USA.
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