Role of specific response elements of the c-fos promoter and involvement of intermediate transcription factor(s) in the induction of Sertoli cell differentiation (transferrin promoter activation) by the testicular paracrine factor PModS.
A mesenchymal-epithelial cell interaction exists in the testis between the Sertoli cells that form the seminiferous tubule and the mesenchymal-derived peritubular myoid cells that surround the tubule. Analysis of the mesenchymal-epithelial interactions between these cells revealed the local production of a mesenchymal factor, PModS. PModS modulates the differentiated functions of Sertoli cells in vitro, including stimulation of the iron-binding protein transferrin (Tf). Previous results have indicated that PModS-induced Tf gene expression involves the activation of immediate early genes. One of the immediate early genes was identified as c-fos. The importance of c-fos was demonstrated in the current study when a c-fos antisense oligonucleotide was found to inhibit the ability of PModS to induce the expression of a Tf promoter-chloramphenicol acetyltransferase (CAT) construct. The regulation of c-fos by PModS was investigated with various CAT constructs containing segments of the c-fos promoter, such as the serum response element (SRE), sis-inducible element (SIE), cAMP response element (CRE), and phorbol ester/TPA response element (TRE), transfected into cultured Sertoli cells. PModS has no effect on cAMP response element-CAT or TRE-CAT, suggesting that PModS does not act through stimulation of cAMP and protein kinase C pathways. PModS was found to activate the c-fos SRE-CAT construct and the SIE-CAT construct. A construct containing both SIE and SRE was stimulated to the same degree as either element alone. Gel mobility shift assays using nuclear extracts from PModS-stimulated Sertoli cells and a radiolabeled SRE oligonucleotide resulted in retarded mobility of a DNA-protein complex. A gel shift with a SRE oligonucleotide containing an ETS domain resulted in a unique shift only detected in PModS stimulated cells. PModS also promoted a gel shift with the SIE that is adjacent to the SRE on the c-fos promoter. The data imply that PModS can activate the c-fos promoter through the SRE and SIE. PModS caused a labeled activating protein 1 (AP1) oligonucleotide to form a DNA-protein complex, indicating activation of the c-fos gene and binding of the c-fos/jun complex. To study the downstream regulation of Sertoli cell differentiation, Tf gene expression was examined. CAT constructs containing deletion mutants of a 3-kilobase (kb) mouse Tf promoter were used. When transfected into Sertoli cells the 581-base pair Tf minimal promoter had only a slight response to PModS, but was activated by FSH. The 2.6-kb Tf promoter construct responded to PModS.(ABSTRACT TRUNCATED AT 400 WORDS)
Whaley
P D
PD
Reproductive Endocrinology Center, University of California, San Francisco 94143-0556, USA.
Chaudhary
J
J
Cupp
A
A
Skinner
M K
MK
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002454:Cell Differentiation / Q000187:drug effects*; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D016762:Genes, fos / Q000187:drug effects; D008297:Male; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D051381:Rats; D012708:Sertoli Cells / Q000166:cytology*; D013735:Testicular Hormones / Q000494:pharmacology*; D014157:Transcription Factors / Q000494:pharmacology*; D014168:Transferrin / Q000235:genetics
C054951:P-Mod-S paracrine factor; D013735:Testicular Hormones; D014157:Transcription Factors; D014168:Transferrin; D004247:DNA; D015500:Chloramphenicol O-Acetyltransferase