Genetic Effects--Disease-Causing Mutations/LH Receptor

促黄体生成素受体失活突变的生化效应 梁，熊，吴，Rennert，陈与施泰因巴赫，Fechner合作。 黄体生成素/绒毛膜促性腺激素受体（LHR）在性腺发育和生殖生理中起着关键作用。在睾丸中，LHR激活导致睾酮的产生。LHR的失活导致睾酮的产生减少，并导致Leydig细胞发育不全（LCH）的高促性腺激素性功能减退症，这是一种男性假两性畸形。一个新的错义突变A340 T与LCH患者中发现的Ile-114取代苯丙氨酸，这影响了一个富含亮氨酸的重复序列（LRR）的细胞外结构域的hLHR。在瞬时表达研究中，突变型受体在hCG刺激后未能触发cAMP产生。这种突变不影响受体的运输，如通过对该受体的绿色荧光蛋白的荧光显微镜研究所揭示的。它影响受体与激素的结合。生成LRR的计算机模型以研究这种突变的影响。该模型清楚地证明了突变的构象效应。这一发现可以扩展到解释突变对其他具有LRR的蛋白质的生物活性的影响。 促黄体生成素/绒毛膜促性腺激素受体在神经系统中的新功能 孟、雷纳特、陈 携带激活突变的LHR个体发展为家族性男性限制性早熟（FMPP），并且经常表现出行为问题。原因尚不清楚。FMPP患者的行为问题可能与LHR突变表达引起的脑细胞功能障碍有关。最近发现LHR在包括神经系统在内的几种非性腺组织中表达。LHR在大脑中的表达水平是发育调节的。在成年大鼠中，在特定脑区的一些神经元、所有四个脑室的室管膜细胞和脉络丛中检测到LHR表达。LHR在这些细胞中的功能尚不清楚。本研究的目的是研究LHR在大脑中的功能活动。将带有激活突变Asp 578 His（H突变）的人LHR cDNA（hLHR）插入pIRES 2 EGFP中，在CMV启动子的控制下，用表达构建体转染大鼠神经元细胞系PC 12。用突变的hLHR转染导致10.8 +/-1.8%的细胞中神经突生长的起始，这显著高于用单独的载体转染的细胞（3.9 +/-0.8%）或用野生型hLHR转染的细胞（3.4 +/-0.7%）中观察到的。在携带神经突的细胞中，与用载体（15.5%）或野生型hLHR（19.2%）转染的细胞相比，用突变的LHR转染的细胞具有显著更高比例的神经突长于2x细胞体的细胞（41.7%）。这些结果提示hLHR在脑内可能具有神经营养作用。进一步的实验正在进行中，以阐明LHR诱导神经元分化的机制。

Biochemical Effect of An Inactivating Mutation of the Luteinizing Hormone Receptor Leung, Bear, Wu, Rennert, Chan in collaboration with Steinbach, Fechner. Luteinizing hormone/Chorionic gonadotropin Receptor (LHR) plays a key role in the development of the gonad and in reproductive physiology. In the testis, LHR activation leads to the production of testosterone. Inactivation of the LHR results in reduced production of testosterone and causes hypergonadotrophic hypogonadism in Leydig Cell Hypoplasia (LCH), a form of male pseudohermaphroditism. A novel missense mutation A340T identified in a patient with LCH resulted in substitution of Ile-114 by Phe, which affects one of the Leucine-rich repeats (LRR) in the extracellular domain of the hLHR. The mutant receptor failed to trigger cAMP production upon hCG stimulation in transient expression studies. This mutation did not affect trafficking of the receptor as revealed by fluorescent microscopic study of the green fluorescent protein of this receptor. It affected binding of the hormone by the receptor. A computer model of the LRR was generated to study the effect of this mutation. The model clearly demonstrated the conformational effect of the mutation. This finding may be extended to explain the impact of mutations on the biological activity of other proteins with LRRs. Novel Function of Luteinizing hormone/Chorionic Gonadotropin Receptor (LHR) in the Nervous System Meng, Rennert, Chan Individuals with LHR carrying activating mutations develop familial male-limited precocious puberty (FMPP) and are often shown to have behavioral problems. The cause is unclear. The behavioral problem of FMPP patients may be related to the dysfunction of brain cells caused by the expression of the mutated LHR. Recently LHR was shown to express in several non-gonad tissues including the nervous system. LHR expression level in the brain is developmentally regulated. In the adult rat LHR expression was detected in some neurons of specific brain regions, the ependymal cells of all four ventricles and the choroid plexus. The function of LHR in these cells is unknown. The goal of this study is to investigate the functional activity of LHR in the brain. The rat neuronal cell line, PC12 was transfected with an expression construct with the human LHR cDNA (hLHR) carrying an activating mutation, Asp578His (H mutation), inserted into pIRES2EGFP under the control of CMV promoter. Transfection with the mutated hLHR led to an initiation of neurite outgrowth in 10.8 +/- 1.8 % of cells which was significantly higher than that observed in cells transfected with the vector alone (3.9 +/- 0.8%) or transfected with the wild type hLHR (3.4 +/- 0.7%). Among the neurite-bearing cells, those transfected with the mutated LHR had a significantly higher proportion of cells that had neurites longer than 2x cell body (41.7%) as compared to those transfected with vector (15.5%) or wild type hLHR (19.2%). These results suggested the possible neurotrophic effects of hLHR in the brain. Further experiments are underway to elucidate the mechanism of LHR induced neuronal differentiation.