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LHCGR Expression During FSH-Induced Follicle Growth is Negatively Regulated by Eukaryotic Initiation Factor 5A. Gulappa T et al. (2017) We have shown that the transient changes in the expression of LHCGR mRNA during the ovarian cycle occurs, at least in part, through a post-transcriptional mechanism involving an LHCGR mRNA binding protein (LRBP). Eukaryotic initiation factor 5A (eIF5A), an LRBP-interacting protein, participates in this process. EIF5A undergoes hypusination, a unique post-translational modification, which is necessary for its functions. The present study examined the role of eIF5A in FSH-induced LHCGR expression during follicular growth. Treatment of primary cultures of rat granulosa cells with FSH and 17β-estradiol showed a time-dependent increase in LHCGR mRNA expression. Conversely, inhibition of endogenous hypusination of eIF5A using GC7, a hypusination inhibitor, showed a greater increase in LHCGR mRNA expression over that produced by FSH and17β-estradiol alone. Further studies were carried out to determine the mechanism by which inhibition of hypusination of eIF5A causes an increase in LHCGR mRNA expression. Since LHCGR expression is negatively regulated by LRBP, the effect of inhibiting hypusination of eIF5A on LRBP expression was examined. The results showed a decrease in the expression of LRBP mRNA and protein when hypusination of eIF5A was inhibited by GC7. Since LRBP has been shown to promote LHCGR mRNA degradation, the present results support the notion that by inhibiting eIF5A hypusination, FSH reduces the expression of LRBP. This increases LHCGR mRNA expression by abrogating the inhibitory action of LRBP.////////////////// Initiation Factor 5A Plays an Essential Role in Luteinizing Hormone Receptor Regulation. Menon B 2014 et al.
Downregulation of LH receptor (LHR) in the ovary by its ligand is mediated by a specific RNA binding protein designated as LHR mRNA binding protein (LRBP), through translational suppression and mRNA degradation. Using yeast two hybrid screens, we previously identified eukaryotic initiation factor 5A (eIF5A) as one of the proteins that interacts with LRBP during LHR mRNA downregulation. The present study examined the role of eIF5A and its hypusination in the context of LHR mRNA downregulation. The association of eIF5A with LRBP or LHR mRNA was determined using immunoprecipitation and RNA Immunoprecipitation (RNA-IP) assays. The results showed that the association of eIF5A with LHR mRNA-LRBP complex increased significantly during downregulation. Furthermore, gel fractionation and hypusination activity assay both showed increased hypusination of eIF5A during LHR mRNA downregulation. Abolishment of hypusination by pretreatment with the chemical inhibitor, GC7 prevented the association of eIF5A with LHR mRNA and LRBP. Inhibition of hypusination also reduced the extent of ligand-induced downregulation of LHR mRNA as well as the expression of functional LH receptors assessed by real time PCR and (125)I-hCG binding assays, respectively. Loss of hCG-mediated downstream signaling during LH receptor downregulation was also restored by inhibition of hypusination of eIF5A. Thus, the present study, for the first time reveals the crucial role of eIF5A and its hypusination in the regulation of LH receptor expression in the ovary.
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