Conformational Change in the Ligand-Binding Pocket via a KISS1R Mutation (P147L) Leads to Isolated Gonadotropin-Releasing Hormone Deficiency.
Conformational Change in the Ligand-Binding Pocket via a KISS1R Mutation (P147L) Leads to Isolated Gonadotropin-Releasing Hormone Deficiency.
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DOI：
10.1210/js.2017-00277
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发表时间：
2017-10-01
影响因子：
4.1
通讯作者：
Nakazato M
中科院分区：
文献类型：
--
作者：
Shimizu K;Yonekawa T;Yoshida M;Miyazato M;Miura A;Sakoda H;Yamaguchi H;Nakazato M
Kisspeptin受体（KISS1R）在下丘脑促性腺激素释放激素神经元中表达，与青春期发病和生殖功能有关。KISS1R突变仍然是先天性促性腺功能低下（CHH）的罕见原因。本研究的目的是确定患者CHH的遗传原因，并对KISS1R突变进行功能表征。患者是一名47岁的日本男子，他的父母是表亲。由于正常CHH，他缺乏第二性征。对该患者的KISS1R基因外显子片段进行突变筛选。使用表达KISS1R突变体的HEK293细胞进行功能分析。分子动力学模拟比较了配体-KISS1R突变复合体与野生型KISS1R变异复合体。在KISS1R中发现了一个纯合突变（c.440C>T, p.P147L）。在重组表达系统中，P147L突变既不影响受体表达水平，也不影响亚细胞定位。细胞内钙测量和细胞介电光谱表明，P147L突变对受体功能的损害程度比先前报道的L148S突变更严重。受体-配体结合试验表明，P147L突变导致配体结合亲和力的大量丧失。分子动力学模拟显示，P147L突变减少了扩展的配体结合口袋中配体-受体复合物的接触表面积。我们在KISS1R中发现了一个与CHH相关的功能缺失突变。我们的研究结果表明，P147L突变导致严重的表型和功能损伤，这是由于配体结合口袋扩大导致配体结合亲和力的丧失。我们发现了一种新的KISS1R突变（P147L），并证明突变受体的配体结合袋的构象变化导致先天性促性腺功能低下。
Kisspeptin receptor (KISS1R) is expressed in hypothalamic gonadotropin-releasing hormone neurons and responsible for pubertal onset and reproductive functions. KISS1R mutations remain a rare cause of congenital hypogonadotropic hypogonadism (CHH). The aim of this study was to identify the genetic cause of CHH in a patient and to functionally characterize a KISS1R mutation. The patient was a 47-year-old Japanese man whose parents were first cousins. He lacked secondary sexual characteristics owing to normosmic CHH. Exon segments for the KISS1R gene in this patient were screened for mutations. Functional analyses were performed using HEK293 cells expressing KISS1R mutants. Molecular dynamics simulations were performed to compare the ligand-KISS1R mutant complex with those of wild-type KISS1R variants. A homozygous mutation (c.440C>T, p.P147L) in KISS1R was identified. The P147L mutation did not affect either receptor expression level or subcellular localization in the recombinant expression system. Intracellular calcium measurements and cellular dielectric spectroscopy demonstrated that the P147L mutation impaired receptor function to an extent more severe than that of a previously reported L148S mutation. A receptor-ligand binding assay showed the P147L mutation causes a substantial loss of ligand-binding affinity. Molecular dynamics simulations revealed the P147L mutation decreases the contact surface area of the ligand-receptor complex in an expanded ligand-binding pocket. We identified a loss-of-function mutation in KISS1R associated with CHH. Our results demonstrated that the P147L mutation causes a severe phenotype and functional impairment resulting from the loss of ligand-binding affinity due to an expanded ligand-binding pocket. We identified a novel KISS1R mutation (P147L) and demonstrated that a conformational change in the ligand-binding pocket of the mutant receptor leads to congenital hypogonadotropic hypogonadism.