Helle K. Knutsen

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 α, β, δ, and ζ messenger RNA (mRNA) and protein in Sertoli cell primary cultures. Treatment with FSH or 8-CPTcAMP strongly induced C/EBP β 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 β mRNA was induced approximately 50-fold, C/EBP δ mRNA was induced 5- to 8-fold by cAMP in Sertoli cells. Messenger RNA for C/EBP β and δ 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 β, δ, and ζ by cAMP. Electrophoretic mobility shift analysis of nuclear proteins from cAMP-treated Sertoli cells using a C/EBP consensus oligonucleotide and antibodies revealed speci...

Inhibitors of RNA and protein synthesis stabilize messenger RNA for the RIIβ subunit of protein kinase a in different cellular compartments

Messenger RNA for RII beta is transiently induced (greater than 50-fold) by cAMP analogs in primary cultures of rat Sertoli cells. The induction is dependent on protein synthesis. We have previously shown that mRNA for RII beta is stabilized by cAMP, as well as inhibitors of transcription and translation. This indicated that rapid degradation of RII beta mRNA involved a protein with a rapid turnover and its corresponding mRNA. The two RNA synthesis inhibitors used in the present study stabilized both nuclear and cytoplasmic RII beta mRNA, whereas inhibition of protein synthesis stabilized RII beta mRNA in the cytoplasm only. These results indicate that only cytoplasmic degradation of RII beta mRNA is dependent on a protein with high turnover. In contrast, nuclear degradation appears to be dependent on an RNA with a short half-life, not involving protein synthesis.

Half-lives of different sized mRNAs for the PKA subunit Rlα are regulated differently in response to inhibition of transcription and translation

The RI alpha mRNA level is induced 3-5 times by FSH or cAMP analogs in primary cultures of rat Sertoli cells. In rat tissues, the RI alpha gene gives rise to three different mRNAs of different size: 3.2, 2.9 and 1.7 kb. In the present study we report that the 1.7 kb transcript has a shorter half-life than the two other mRNAs. In cells which had been pre-stimulated with a cAMP analog, inhibition of transcription stabilizes the two larger, but not the smaller sized RI alpha mRNA. However, in contrast, inhibition of protein synthesis stabilizes all the RI alpha mRNAs. Thus, degradation of various mRNAs coding for the same protein reveals different dependencies on transcription and translation.