EPIGENOMIC RELATIONSHIP TO A SEX DIFFERENCE IN HYPOTHALAMIC LHRH MRNA EXPRESSION

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 目的：探讨促肾上腺皮质激素释放激素(LHRH)基因表达是否存在性别差异，以及其他已知影响LHRH神经元活性的神经系统。 黄体生成素释放激素(LHRH)是生殖功能的关键调节因子。成年恒河猴LHRH基因在下丘脑的表达受性腺类固醇激素的调节，雌性主要是雌激素，雄性主要是雄激素。尽管存在这种差异，但目前还没有关于性别特异性信号如何调控LHRH基因表达的报道。 从性腺完整的成年雄性恒河猴(n=5，11.0±1.7岁)和雌性恒河猴(n=5，11.2±2.0岁)的下丘脑内侧基底核(MBH)和视前区(POA)收集RNA和DNA。雌性猴子排卵周期的阶段是未知的，因为它们是从组织分配程序中获得的。定量聚合酶链式反应发现，男性MBH中LHRH基因表达水平显著低于女性(p0.05)，而POA中LHRH基因表达水平无明显差异。此外，我们还发现GAD65、GAD67、NPY、KISS-1和GPR54基因在这两个地区的表达没有性别差异。由于灵长类下丘脑LHRH神经分泌系统不存在永久性的性别分化，即卵巢移植可诱导去成年雄性猕猴的黄体生成素高峰(Norman and Spies Endocinology 118：2608-2610,1986)，因此LHRH基因表达的性别差异可能是由表观遗传机制调节的。在第二个实验中，我们调查了CpG甲基化是否在性别之间存在差异。CpG甲基化是DNA的一种共价修饰，影响转录因子结合和基因表达。初步调查显示，在LHRH基因的5‘侧翼区，男性比女性表现出更高的CpG甲基化的趋势(p0.1)。该甲基化位于恒河猴LHRH基因转录起始点的CpG富集区-2100/-1700。总之，这些结果表明，恒河猴LHRH mRNA水平存在显著的性别差异，这可能与CpG甲基化的差异有关。结论：表观基因组修饰是灵长类LHRH基因表达复杂编排的影响因素。 本研究使用了WNPRC动物服务(病理学服务)。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Objective: To investigate whether sex differences exist in gene expression of LHRH as well as other neuronal systems known to influence LHRH neuronal activity. Luteinizing-hormone releasing hormone (LHRH) is a key regulator of reproductive function. Adult rhesus monkey (macaca mulatta) LHRH gene expression in the hypothalamus is regulated by gonadal steroids, primarily estrogens in females and androgens in males. Despite this distinction, there is no report of how sex-specific signals regulate LHRH gene expression. RNA and DNA were collected from the medial basal hypothalamus (MBH) and preoptic area (POA) of gonadally intact adult male (n=5, 11.0¿1.7 years of age) and female (n=5, 11.2¿2.0 years of age) rhesus monkeys. The phases of ovulatory cycle in the female monkeys were unknown, as they were obtained from the tissue distribution program. Using quantitative PCR, we found that LHRH mRNA levels in the MBH are significantly lower in males compared to females (p0.05), but there was no difference in the POA. Moreover, we found that there were no sex differences in GAD65, GAD67, NPY, KiSS-1, and GPR54 mRNA in either area. Because there is no permanent sexual differentiation in the primate hypothalamic LHRH neurosecretory system, i.e., LH surges could be induced by ovarian transplantation in orchidectomized adult male monkeys (Norman and Spies Endocrinology 118:2608-2610, 1986), it is possible that an epigenetic mechanism regulates the sex difference in LHRH gene expression. In the second experiment we investigated if CpG methylation, a covalent modification of DNA that influences transcription factor binding and gene expression, was different between the sexes. Preliminary investigations revealed a trend (p0.1) with males exhibiting higher CpG methylation than females in the 5' flanking region of the LHRH gene. This methylation is in a CpG rich region -2100/-1700 relative to the transcription start site in the rhesus LHRH gene. Collectively, these results indicate that there is a significant sex difference in LHRH mRNA levels in rhesus monkeys and that this may be related to a difference in CpG methylation. It is concluded that epigenomic modifications are factors in the complex orchestration of LHRH gene expression in primates. This research used WNPRC Animal Services (Pathology Services).