Regulation of luteinizing hormone receptor expression by an RNA binding protein: Role of ERK signaling

A specific luteinizing hormone receptor (LHR) mRNA binding protein (LRBP) has been identified and purified. This LH receptor mRNA binding protein selectively binds to the polypyrimidine rich bipartite sequence in the coding region of the LHR mRNA and accelerates its degradation. In response to preovulatory LH surge, the LH receptor expression in the ovary undergoes downregulation by accelerated degradation of LH receptor mRNA through the involvement of this RNA binding protein. Here we describe the intracellular mechanism triggered by LH/hCG (human chorionic gonadotropin) that leads to the regulated degradation of LH receptor mRNA. Downregulation of LH receptor mRNA was induced by treatment of cultured human granulosa cells with 10 IU of hCG. Activation of downstream target, extracellular signal-regulated protein kinase 1 and 2 (ERK 1/2) showed an increase within five min and sustained up to 1 h. Confocal analysis showed that ERK1/2 translocates to the nucleus after 15 min of hCG treatment. This leads to an increase in LRBP expression which then causes downregulation of LH receptor mRNA by accelerating its degradation. Treatment with UO126 or transfection with ERK specific siRNA (small interfering RNA) resulted in the abolishment of ERK activation as well as LHR mRNA downregulation. RNA electrophoretic mobility gel shift assay of the cytosolic fractions showed that hCG-induced increase in the LH receptor mRNA binding activity was also abrogated by these treatments. These results show that LH/hCG-induced LH receptor mRNA downregulation is initiated by the activation of ERK1/2 pathway by regulating the expression and activity of LH receptor mRNA binding activity

Regulation of Sterol Regulatory Element-Binding Transcription Factor 1a by Human Chorionic Gonadotropin and Insulin in Cultured Rat Theca-Interstitial Cells1

Theca-interstitial (T-I) cells of the ovary synthesize androgens in response to luteinizing hormone (LH). In pathological conditions such as polycystic ovarian syndrome, T-I cells are hyperactive in androgen production in response to LH and insulin. Because cholesterol is an essential substrate for androgen production, we examined the effect of human chorionic gonadotropin (hCG) and insulin on signaling pathways that are known to increase cholesterol accumulation in steroidogenic cells. Specifically, the effect of hCG and insulin on sterol regulatory element-binding transcription factor 1a (SREBF1a) required for cholesterol biosynthesis and uptake was examined. Primary cultures of T-I cells isolated from 25-day-old rat ovaries responded to hCG and insulin to increase the active/processed form of SREBF1a. The hCG and insulin significantly reduced insulin-induced gene 1 (INSIG1) protein, a negative regulator of SREBF processing. Furthermore, an increase in the expression of selected SREBF target genes, 3-hydroxy-3-methylglutaryl-coenzyme A reductase (Hmgcr) and mevalonate kinase (Mvk), was also observed. Protein kinase A (PRKA) inhibitor completely abolished the hCG-induced increase in SREBF1a, while increasing INSIG1. Although the hCG-induced depletion of total and free cholesterol was abolished by aminoglutethimide, the stimulatory effect on SREBF1a was not totally suppressed. Treatment with 25-hydroxycholesterol abrogated the effect of hCG on SREBF1a. Inhibition of the phosphatidylinositol 3-kinase pathway did not block the insulin-induced increase in SREBF1a, whereas mitogen-activated protein kinase inhibition reduced the insulin response. These results suggest that the increased androgen biosynthesis by T-I cells in response to hCG and insulin is regulated, at least in part, by increasing the expression of sterol response element-responsive genes by increasing SREBF1a