[Stimulation of LH gene expression by GnRH. Role of protein kinases A and C].

Using anterior pituitary cells in culture, incubated in the presence of [35S] Met, we have previously demonstrated that GnRH stimulates the synthesis of LH polypeptide chains, an effect which was reproduced in a non additive manner by direct activation of protein kinases A and C (by cyclic AMP or diacylglycerol analogues, respectively), and abolished by actinomycin D. The respective roles of GnRH and protein kinases A and C in the stimulation of LH subunit gene expression was examined in the present study by evaluating the effects on the cellular levels of alpha and LH beta mRNAs of optimal concentrations of Buserelin (0.1 nM) an undegradable GnRH analogue, cholera toxin (6 nM) a cyclic AMP (AMPc) generator and 12-O-tetradecanoyl phorbol 13-acetate (TPA) a diacylglycerol analogue. The specific mRNAs were quantified by densitometry analysis of Northern blot hybridization using 32P-labeled single strand cDNAs. We observed TPA, like Buserelin, increased LH mRNA levels in a very similar manner, the alpha mRNA increasing 1.8 - 2.5 fold and the LH beta mRNA 1.4 - 1.7 fold after 5 h culture and 3.0 - 3.5 fold and 2.2 - 2.4 fold respectively, after 24 h. In the presence of cholera toxin, the changes were more rapid, the highest values being reached at 5 h (8.6 fold increase for alpha and 4.0 fold for LH beta mRNA). LH, radioimmunoassayed in parallel in the cell medium, increased 5.9 fold after 5 h and 7.1 fold after 24 h culture in the presence of Buserelin, and 2.9 fold and 5.4 fold, respectively, in the presence of TPA.(ABSTRACT TRUNCATED AT 250 WORDS)