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Rescue of Function of Mutant Luteinising Hormone Receptors with Deficiencies in Cell Surface Expression, Hormone Binding, and Hormone Signalling.
Neuroendocrinology ( IF 3.2 ) Pub Date : 2020-04-21 , DOI: 10.1159/000508000
Claire Louise Newton 1, 2, 3 , Ross Calley Anderson 4, 5 , Annika Kreuchwig 6 , Gerd Krause 6 , Arieh Anthony Katz 7 , Robert Peter Millar 4, 8, 9, 10
Affiliation  

INTRODUCTION G protein-coupled receptor (GPCR) mutations are implicated in many diseases. Most inactivating mutations cause receptor misfolding and prevent trafficking to the plasma membrane. Pharmacological chaperones can "rescue" cell surface expression of such mutants, presumably by stabilising correct folding of the nascent protein. OBJECTIVE Here we examine the scope of intracellularly retained luteinising hormone receptor (LHR) mutants that can be "rescued" by the pharmacological chaperone LHR-Chap, and whether this allosteric agonist can also restore the function of mutant LHRs with deficiencies in hormone binding or hormone-induced signalling. METHODS Mutant LHRs were expressed in HEK 293-T cells. Cell surface expression/localisation, hormone binding, and hCG/LHR-Chap signalling were determined by ELISA, radioligand binding, and inositol phosphate accumulation assays, respectively. Molecular modelling predicted LHR-Chap interactions. RESULTS LHR-Chap increased cell surface expression of a subset of retained mutants located in transmembrane helices predicted to be stabilised by LHR-Chap binding. For 3 (T4613.47I, L5024.61P, and S6167.46Y) hCG-responsiveness was increased following treatment. LHRs with mutations in the hormone-binding site (C131ECDR and I152ECDT) or in the hinge region (E354HingeK) had good cell surface expression but poor response to hormone stimulation, yet were responsive to allosteric activation by LHR-Chap. CONCLUSIONS LHR-Chap, in addition to rescuing cell surface expression of intracellularly retained LHR mutants, can rescue function in mutant receptors with binding and signalling deficiencies that have normal cell surface expression. This demonstration of rescue of multiple elements of LHR dysfunction arising from inactivating mutations offers exceptional potential for treating patients with diseases arising from GPCR mutations in general.

中文翻译:

细胞表面表达、激素结合和激素信号传导缺陷的突变促黄体激素受体功能的拯救。

引言 G 蛋白偶联受体 (GPCR) 突变与许多疾病有关。大多数失活突变会导致受体错误折叠并阻止向质膜运输。药理学伴侣可以“拯救”这些突变体的细胞表面表达,大概是通过稳定新生蛋白质的正确折叠。目的 在这里,我们检查了可以被药理学伴侣 LHR-Chap“拯救”的细胞内保留的促黄体生成素受体 (LHR) 突变体的范围,以及这种变构激动剂是否也可以恢复具有激素结合或激素缺陷的突变体 LHRs 的功能-诱导信号。方法 突变的 LHRs 在 HEK 293-T 细胞中表达。细胞表面表达/定位、激素结合和 hCG/LHR-Chap 信号通过 ELISA 确定,放射性配体结合和肌醇磷酸积累测定,分别。分子建模预测 LHR-Chap 相互作用。结果 LHR-Chap 增加了位于跨膜螺旋中的保留突变体子集的细胞表面表达,预测通过 LHR-Chap 结合而稳定。对于 3(T4613.47I、L5024.61P 和 S6167.46Y),治疗后 hCG 反应性增加。在激素结合位点(C131ECDR 和 I152ECDT)或铰链区(E354HingeK)突变的 LHR 具有良好的细胞表面表达,但对激素刺激的反应较差,但对 LHR-Chap 的变构激活有反应。结论 LHR-Chap 除了挽救细胞内保留的 LHR 突变体的细胞表面表达外,可以挽救具有正常细胞表面表达的结合和信号传导缺陷的突变受体的功能。这种拯救由失活突变引起的 LHR 功能障碍的多个因素的证明为治疗患有一般 GPCR 突变引起的疾病的患者提供了非凡的潜力。
更新日期:2020-04-21
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