Follicle-stimulating hormone increases the turnover of G-protein αi-1- and αi-2-subunit messenger RNA in Sertoli cells by a mechanism that is independent of protein synthesis

Abstract

We have shown previously that FSH differentially regulated G-protein αi-subunit mRNA levels in primary cultures of rat Sertoli cells. The mechanism by which FSH regulated the steady state levels of these mRNAs was investigated by assessing the effect of FSH on G-protein α-subunit mRNA stability and the potential involvement of newly synthesized proteins. The half-life (t1/2) of α-subunit mRNA was determined by inhibiting Sertoli cell transcription with actinomycin-D and quantifying α-subunit mRNA levels by Northern blot analyses. Transcripts for αi-2 and αs were extremely stable, possessing t1/2 of 44 and 51 h, respectively. In contrast, the t1/2 of αi-3 mRNA was only 3.6 h. Turnover of αi-1 mRNA occurred as a two-phase decay, with an initial t1/2fast of 0.8 h, followed by a second phase with t1/2slow of 11.1 h. Treatment of Sertoli cells with FSH in the presence of actinomycin-D destabilized the αi-1 mRNA to a single phase decay with a t1/2 of 5.7 h. FSH also decreased the stability of the αi-2 mRNA from a t1/2 of 51 h to a t1/2fast of 16.5 h. These effects of FSH on αi-1 and αi-2 mRNA stability may contribute to the ability of FSH to decrease steady state αi-1 and αi-2 mRNA levels. The effect of FSH on αi-3 mRNA stability was not significant, suggesting an alternative mechanism regulating the FSH-mediated increase of αi-3 mRNA levels. Inhibition of protein synthesis by cycloheximide significantly increased steady state levels of α-subunit mRNA, but did not influence FSH-induced alterations in α-subunit mRNA levels. These data indicate that FSH did not require newly translated proteins to mediate its effects. The FSH-induced decay of αi-1 and αi-2 was inhibited partially by the addition of actinomycin-D, suggesting that normal turnover of these mRNA is mediated by labile destabilizing proteins. We propose that FSH regulates αi-1 and αi-2 mRNA levels at least in part by increasing G-protein mRNA turnover, which may require modification of preexisting proteins.