Transcriptional regulation of chorionic gonadotropin alpha- and beta-subunit gene expression by 8-bromo-adenosine 3',5'-monophosphate.
In placental explants and cell lines, cAMP stimulates the biosynthesis of CG alpha- and beta-subunits and produces marked stimulation of CG secretion. We investigated the effect of a cell-permeable analog, 8-bromo-cAMP (8-Br-cAMP), on CG alpha- and beta-subunit gene expression in the JEG-3 placental cell line. RNA blot hybridization analysis demonstrated that 8-Br-cAMP increased CG alpha and beta mRNA levels 15- to 30-fold, consistent with 8-Br-cAMP regulation of CG biosynthesis at a pretranslational level. Transcription rates for CG alpha- and beta-subunit genes were determined in the absence and presence of 8-Br-cAMP using nuclear run-on assays. The basal transcription rates for the alpha- and beta-subunit genes were similar and increased 4- to 6-fold after treatment with 8-Br-cAMP. Expression of chimeric genes, consisting of alpha gene [1.5 kilobase (Kb)] or CG beta gene (0.3 Kb) 5'-flanking sequences ligated to the coding sequence of a reporter enzyme chloramphenicol acetyl transferase (CAT), was used to analyze the mechanism of 8-Br-cAMP stimulation of gene transcription. A 1.5-kb segment of the alpha- gene 5'-flanking sequence was expressed efficiently in JEG-3 cells and contains both cell-specific enhancers and cAMP response elements. Basal expression of the CG beta CAT reporter gene was not observed in either JEG-3 cells or HeLa cells, and expression was not stimulated by 8-Br-cAMP. The regulatory response elements of the CG beta gene must reside in sequences outside the 0.3-kb 5'-flanking region of the CG beta gene.
Jameson
J L
JL
Jaffe
R C
RC
Gleason
S L
SL
Habener
J F
JF
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology*; D000123:Acetyltransferases / Q000235:genetics; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase; D006063:Chorionic Gonadotropin / Q000235:genetics*; D005786:Gene Expression Regulation / Q000187:drug effects; D006367:HeLa Cells; D006801:Humans; D012333:RNA, Messenger / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects
D006063:Chorionic Gonadotropin; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D015124:8-Bromo Cyclic Adenosine Monophosphate; D000123:Acetyltransferases; D015500:Chloramphenicol O-Acetyltransferase