The 5'-flanking region of the rat luteinizing hormone/chorionic gonadotropin receptor gene confers Leydig cell expression and negative regulation of gene transcription by 3',5'-cyclic adenosine monophosphate.
The LH/CG receptor is a G protein-coupled receptor present on gonadal cells whose levels are modulated by a number of hormones, growth factors, and second messenger analogs. With the recently cloned cDNA for the LH/CG receptor, it has been shown that changes in the levels of the cognate mRNA are involved, at least in part, in the observed changes in receptor density. In order to study the transcriptional regulation of the LH/CG receptor we have isolated a 2-kilobase region of the 5'-flanking region of the rat LH/CG receptor gene and subcloned nucleotide -1 (relative to the translational initiation codon) to -1370 into a luciferase reporter plasmid. We show here that this region of the LH/CG receptor gene is able to enhance luciferase activity in MA-10 cells, a line of Leydig tumor cells that normally express LH/CG receptors, as opposed to human kidney 293 cells, which do not. Furthermore, the addition of 8-bromo-cAMP to MA-10 cells, under conditions known to decrease LH/CG receptor numbers and receptor mRNA levels, decreases the relative luciferase activity to about 26% of control. This decrease in reporter gene activity is severely blunted in a subclone of MA-10 cells with a cAMP-resistant phenotype. Our studies show, for the first time, that sequence(s) present with 1370 base pairs of the translational start site of the rat LH/CG receptor gene are sufficient for conferring expression of this gene in Leydig cells and for the negative modulation of LH/CG receptor gene transcription by high concentrations of cAMP.
Wang
H
H
Department of Physiology, University of Iowa College of Medicine, Iowa City 52242.
Nelson
S
S
Ascoli
M
M
Segaloff
D L
DL
D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D003001:Cloning, Molecular; D000242:Cyclic AMP / Q000502:physiology*; D005786:Gene Expression Regulation / Q000502:physiology*; D007985:Leydig Cells / Q000378:metabolism; D008156:Luciferases / Q000235:genetics; D008297:Male; D008969:Molecular Sequence Data; D051381:Rats; D011974:Receptors, LH / Q000235:genetics*; D012045:Regulatory Sequences, Nucleic Acid / Q000502:physiology*; D014158:Transcription, Genetic / Q000502:physiology*
D011974:Receptors, LH; D000242:Cyclic AMP; D008156:Luciferases