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Isoform-Specific Regulation of the CCAAT/Enhancer-Binding Protein Family of Transcription Factors by 3',5'-Cyclic Adenosine Monophosphate in Sertoli Cells¹

Institute of Medical Biochemistry (L.M.G., M.K.D., K.A.T., K.T., H.K.K.), University of Oslo, N-0317 Oslo, Norway; Institutes of Molecular Biology (S.E.) and Physiology (L.H.), University of Gøteborg, S-41390 Gøteborg, Sweden

Address all correspondence and requests for reprints to: Line M. Grønning, M.Sc., Institute of Medical Biochemistry, University of Oslo, P.O. Box 1112 Blindern, N-0317 Oslo, Norway. E-mail: l.m.gronning{at}basalmed.uio.no

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 , ß, , and messenger RNA (mRNA) and protein in Sertoli cell primary cultures. Treatment with FSH or 8-CPTcAMP strongly induced C/EBP ß mRNA above basal levels with rapid and transient kinetics in Sertoli cell primary cultures as well as in whole testes from hypophysectomized rats. Whereas C/EBP ß mRNA was induced approximately 50-fold, C/EBP mRNA was induced 5- to 8-fold by cAMP in Sertoli cells. Messenger RNA for C/EBP ß and were induced by inhibition of protein synthesis with cycloheximide and cycloheximide acted synergistically with cAMP. Immunoblots with C/EBP antibodies demonstrated a strong induction of C/EBP ß, , and by cAMP. Electrophoretic mobility shift analysis of nuclear proteins from cAMP-treated Sertoli cells using a C/EBP consensus oligonucleotide and antibodies revealed specific binding of C/EBP/DNA complexes, the majority of which were supershifted by C/EBP ß antibody. Transfections of Sertoli cells with a C/EBP reporter construct showed approximately 3-fold induction of reporter gene 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 transfections of Sertoli cells. Furthermore, C/EBP ß expression increased the activities of two promoters known to be cAMP-responsive in Sertoli cells. Thus, the early induction of C/EBP isoforms by cAMP may play a role in FSH-dependent regulation of late response genes in Sertoli cells.

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