Follicle stimulating hormone (FSH) stimulates transferrin gene transcription in rat Sertoli cells: cis and trans-acting elements involved in FSH action via cyclic adenosine 3',5'-monophosphate on the transferrin gene.
FSH is a major regulator of transferrin (Tf) production in the testis. FSH effects on Sertoli cell Tf production are believed to be mediated, at least in part, via cAMP second messenger system. Previously, it has been shown that FSH and (Bu)2cAMP stimulate Tf mRNA levels. This study examines the effect of cAMP and FSH on Tf gene transcription using run-on assays. These data demonstrate rapid induction of Tf gene by (Bu)2cAMP (2.3-fold) and FSH (2.8-fold) within 30 min and 2 h, respectively. Furthermore, the ability of (Bu)2cAMP and FSH to drive the transcription of chimeric constructs containing a 0.6-kilobase segment of the 5'-regulatory region of the human Tf gene coupled to a chloramphenicol acetyltransferase (CAT) was examined. Deletion analysis indicated that the sequence -100/-52 base pairs is required for the cAMP-dependent transcription. This sequence shows no homology to that of the consensus cAMP-regulatory element (CRE). However, cotransfection experiments with a CRE-binding protein (CREB) expression vector revealed a basal induction of the Tf transcriptional activity as well as a synergistic activation of CREB and (Bu)2cAMP. Expression of KCREB, a dominant negative mutant form of CREB, completely blocked the cAMP induction of the -100+39Tf-CAT construct. This region contains two functional regions PRI and PRII. Gel shift assay with nuclear proteins from Sertoli cells using the PRII and PRI probes showed that the band shifts formed by PRII were competitive complexes with CRE, and a CREB antiserum retarded the migration of nuclear Sertoli cells proteins. We conclude that CREB is implicated in the FSH regulation on the Tf gene in Sertoli cells.
Suire
S
S
Institut National de la Recherche Agronomique (INRA) Station de Physiologie de la Reproduction des Mammifères Domestiques, Unité de Recherche Associée, Nouzilly, France.
Fontaine
I
I
Guillou
F
F
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D003994:Bucladesine / Q000494:pharmacology; D000242:Cyclic AMP / Q000502:physiology*; D017362:Cyclic AMP Response Element-Binding Protein / Q000235:genetics / Q000502:physiology*; D005640:Follicle Stimulating Hormone / Q000494:pharmacology*; D005813:Genes, Synthetic; D046911:Macromolecular Substances; D008297:Male; D008969:Molecular Sequence Data; D011485:Protein Binding; D051381:Rats; D017208:Rats, Wistar; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid; D015290:Second Messenger Systems; D012708:Sertoli Cells / Q000187:drug effects* / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects*; D014168:Transferrin / Q000096:biosynthesis / Q000235:genetics*
D017362:Cyclic AMP Response Element-Binding Protein; D046911:Macromolecular Substances; D011993:Recombinant Fusion Proteins; D014168:Transferrin; D003994:Bucladesine; D005640:Follicle Stimulating Hormone; D000242:Cyclic AMP