The transcriptional response of the human chorionic gonadotropin beta-subunit gene to cAMP is cycloheximide sensitive and is mediated by cis-acting sequences different from that found in the alpha-subunit gene.
Cyclic AMP stimulates transcription of the genes encoding the alpha- and beta-subunits of human CG (hCG). Although the cis-acting cAMP response element (CRE) of the alpha-subunit gene has been extensively characterized, relatively little is known about the putative response element of the hCG beta gene. Here, we demonstrate that cAMP stimulation of hCG beta gene transcription requires ongoing protein synthesis, whereas cAMP stimulation of alpha-subunit gene transcription does not. These results suggest that cAMP-stimulated transcription of the alpha and hCG beta genes may involve different cis-acting elements and trans-acting factors. Accordingly, we have constructed a series of deletion mutants consisting of fragments of the hCG beta promoter-regulatory region linked to the bacterial chloramphenicol acetyl transferase gene. To potentiate detection of a CRE, we constructed vectors containing the Rous sarcoma virus enhancer to augment transcriptional activity of the hCG beta-promoter. We also used vectors designed to determine whether regions containing a putative CG beta CRE could confer cAMP responsiveness to a heterologous promoter. These constructs were evaluated for activity by transfection into choriocarcinoma (BeWo) cells. Transient expression of these vectors identifies a broad region which renders transcription of the chloramphenicol acetyl transferase gene responsive to cAMP. Surprisingly, division of the hCG beta 5'-flanking and untranslated regions into smaller fragments led to loss of cAMP responsiveness, suggesting that the hCG beta CRE may be comprised of multiple domains dispersed across the proximal 650 base pairs of CG beta 5'-flanking and untranslated sequence.(ABSTRACT TRUNCATED AT 250 WORDS)
Fenstermaker
R A
RA
Department of Pharmacology, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106.
Milsted
A
A
Virgin
J B
JB
Miller
W L
WL
Nilson
J H
JH
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D002467:Cell Nucleus / Q000032:analysis; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D006063:Chorionic Gonadotropin / Q000235:genetics*; D018997:Chorionic Gonadotropin, beta Subunit, Human; D000242:Cyclic AMP / Q000502:physiology; D003513:Cycloheximide / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D005809:Genes, Regulator; D005822:Genetic Vectors; D015292:Glycoprotein Hormones, alpha Subunit / Q000235:genetics*; D009693:Nucleic Acid Hybridization; D010446:Peptide Fragments / Q000235:genetics*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012313:RNA / Q000096:biosynthesis; D014157:Transcription Factors / Q000378:metabolism; D014161:Transduction, Genetic
D006063:Chorionic Gonadotropin; D018997:Chorionic Gonadotropin, beta Subunit, Human; D015292:Glycoprotein Hormones, alpha Subunit; D010446:Peptide Fragments; D011500:Protein Synthesis Inhibitors; D014157:Transcription Factors; D015124:8-Bromo Cyclic Adenosine Monophosphate; D012313:RNA; D003513:Cycloheximide; D000242:Cyclic AMP; D015500:Chloramphenicol O-Acetyltransferase