A Single-Nucleotide Polymorphism in the EAP1 Gene Is Associated with Amenorrhea/Oligomenorrhea in Nonhuman Primates

Abstract

Current evidence suggests that the acquisition of female reproductive capacity and the maintenance of mature reproductive function are related processes transcriptionally regulated by gene networks operating within the neuroendocrine brain. One of these genes, termed enhanced at puberty 1 (EAP1), encodes an upstream regulator of these processes. Selective inhibition of EAP1 expression in discrete regions of the rat and nonhuman primate (NHP) hypothalamus, via targeted delivery of RNA interference, either disrupts (rats) or abolishes (monkeys) reproductive cycles. The striking loss of menstrual cyclicity resulting from knocking down hypothalamic EAP1 expression suggests that diminished EAP1 function may contribute to disorders of the menstrual cycle of neuroendocrine origin. Here we show that a single-nucleotide polymorphism in the 5'-flanking region of EAP1 gene is associated with increased incidence of amenorrhea/oligomenorrhea in NHP. In the presence of the risk allele, binding of the transcription factor mothers against decapentaplegic homolog 3 (SMAD3) to its recognition site contained within the polymorphic sequence in the monkey EAP1 promoter is reduced. The risk allele also diminishes the increase in EAP1 promoter activity elicited by TGFβ1, a peptide that activates a SMAD3/4-mediated signaling pathway to regulate gene transcription. These findings indicate that common genetic variation in the EAP1 locus increases the susceptibility of NHP to loss/disruption of menstrual cyclicity. They also raise the possibility that polymorphisms in EAP1 may increase the risk of functional hypothalamic amenorrhea in humans.

A system biology approach to identify regulatory pathways underlying the neuroendocrine control of female puberty in rats and nonhuman primates

“…Similar injections into the medial basal hypothalamus of rhesus monkeys obliterated menstrual cyclicity (Dissen and others, 2012) indicating that EAP1 is crucial for the ARC to maintain menstrual cyclicity in higher primates. This role is supported by the recent finding that a single nucleotide polymorphism in the 5′-flanking region of the EAP1 gene is associated with reduced EAP1 transcription and oligomenorrhea/amenorrhea in nonhuman primates (Lomniczi and others, 2012a). …”

Neuroendocrine control of the onset of puberty

“…Interestingly, EAP1 is expressed in kisspeptin neurons in the arcuate nucleus of the rat [83]. Moreover, knockdown of EAP1 using a lentivirus approach interrupts menstrual cyclicity in the monkey and a SNP upstream of the EAP1 gene has been associated with irregular menses in this primate [84] [85]. Interrogation of global gene expression in the agonadal male rhesus monkey, however, using a monkey cDNA array failed to reveal an increase expression of EAP1 in the mediobasal hypothalamus during the juvenilepubertal transition when GnRH pulse generation is reactivated (Ojeda and Plant, unpublished observations).…”

Molecular and Gene Network Analysis of Thyroid Transcription Factor 1 <b><i>(TTF1)</i></b> and Enhanced at Puberty <b><i>(EAP1)</i></b> Genes in Patients with GnRH-Dependent Pubertal Disorders

“…Although the EAP1 gene appeared functionally intact in our patients, it is formally possible that inactivating mutations may be found upon analysis of much larger populations of affected individuals or that single nucleotide polymorphisms affecting either the transcriptional regulation of EAP1 or the stability of its encoded mRNA may be discovered. A precedent for this potential outcome has been provided by the recent demonstration that a single-nucleotide polymorphism in the 5′-flanking region of the EAP1 gene is associated with increased incidence of amenorrhea/oligomenorrhea in rhesus monkeys [43]. …”

“…The same holds true for sequence variation affecting the EAP1 gene itself. Three observations provide initial support for this concept: one is the finding that a SNP in the 5′-flanking region of the EAP1 is associated with hypothalamic amenorrhea in non-human primates and with the loss of a SMAD3 site required for TGFβ-dependent activation of EAP1 transcription [43]. The second observation is the presence of a SNP (rs10980926) overlapping a predicted TTF1 site in an intron of ZNF483 ; this SNP was identified as associated with an earlier age at menarche in a meta-analysis of GWAS studies [46].…”