Isoform-specific regulation of the CCAAT/enhancer-binding protein family of transcription factors by 3 ',5 '-cyclic adenosine monophosphate in Sertoli cells

The C/EBP (CCAAT/enhancer-binding protein) family of transcription factors is important for differentiation, lipid biosynthesis, and metabolism. Here, we demonstrate for the first time the presence of C/EBP alpha, beta, delta , and zeta messenger RNA (mRNA) and protein in Sertoli cell primary culture s. Treatment with FSH or 8-CPTcAMP strongly induced C/EBP beta mRNA above b asal levels with rapid and transient kinetics in Sertoli cell primary cultu res as well as in whole testes from hypophysectomized rats. Whereas C/EBP b eta mRNA was induced approximately 50-fold, C/EBP delta mRNA was induced 5- to 8-fold by cAMP in Sertoli cells, Messenger RNA for C/EBP beta and delta were induced by inhibition of protein synthesis with cycloheximide and cyc loheximide acted synergistically with cAMP. Immunoblots with C/EBP antibodi es demonstrated a strong induction of C/EBP beta, delta, and zeta by cAMP. Electrophoretic mobility shift analysis of nuclear proteins from cAMP-treat ed Sertoli cells using a C/EBP consensus oligonucleotide and antibodies rev ealed specific binding of C/EBP/DNA complexes, the majority of which were s upershifted by C/EBP beta antibody. Transfections of Sertoli cells with a C /EBP reporter construct showed approximately 3-fold induction of reporter g ene activity by cAMP. In contrast, the reporter gene vector with a mutated form of the C/EBP binding site, was almost unresponsive to cAMP in transfec tions of Sertoli cells. Furthermore, C/EBP beta expression increased the ac tivities of two promoters known to be cAMP-responsive in Sertoli cells. Thu s, the early induction of C/EBP isoforms by cAMP may play a role in FSH-dep endent regulation of late response genes in Sertoli cells.