Human Molecular Genetics Research Group, Institute of Molecular and Cell Biology, University of Tartu, Riia 23, 51010 Tartu, Estonia.
Mol Hum Reprod. 2012 Aug;18(8):379-90. doi: 10.1093/molehr/gas018. Epub 2012 May 3.
Heterodimeric hCG is one of the key hormones determining early pregnancy success. We have previously identified rare missense mutations in hCGβ genes with potential pathophysiological importance. The present study assessed the impact of these mutations on the structure and function of hCG by applying a combination of in silico (sequence and structure analysis, molecular dynamics) and in vitro (co-immunoprecipitation, immuno- and bioassays) approaches. The carrier status of each mutation was determined for 1086 North-Europeans [655 patients with recurrent miscarriage (RM)/431 healthy controls from Estonia, Finland and Denmark] using PCR-restriction fragment length polymorphism. The mutation CGB5 p.Val56Leu (rs72556325) was identified in a single heterozygous RM patient and caused a structural hindrance in the formation of the hCGα/β dimer. Although the amount of the mutant hCGβ assembled into secreted intact hCG was only 10% compared with the wild-type, a stronger signaling response was triggered upon binding to its receptor, thus compensating the effect of poor dimerization. The mutation CGB8 p.Pro73Arg (rs72556345) was found in five heterozygotes (three RM cases and two control individuals) and was inherited by two of seven studied live born children. The mutation caused ~50% of secreted β-subunits to acquire an alternative conformation, but did not affect its biological activity. For the CGB8 p.Arg8Trp (rs72556341) substitution, the applied in vitro methods revealed no alterations in the assembly of intact hCG as also supported by an in silico analysis. In summary, the accumulated data indicate that only mutations with neutral or mild functional consequences might be tolerated in the major hCGβ genes CGB5 and CGB8.
异二聚体 hCG 是决定早期妊娠成功的关键激素之一。我们之前已经鉴定出 hCGβ 基因中具有潜在病理生理学意义的罕见错义突变。本研究通过应用计算机模拟（序列和结构分析、分子动力学）和体外（共免疫沉淀、免疫和生物测定）方法，评估了这些突变对 hCG 结构和功能的影响。使用 PCR-限制性片段长度多态性，对来自爱沙尼亚、芬兰和丹麦的 655 名复发性流产（RM）/431 名健康对照的 1086 名北欧人（每个突变的携带者状态进行了确定。在单个杂合性 RM 患者中发现了突变 CGB5 p.Val56Leu（rs72556325），导致 hCGα/β二聚体形成的结构障碍。尽管与野生型相比，组装成分泌完整 hCG 的突变 hCGβ 量仅为 10%，但与受体结合时会触发更强的信号反应，从而补偿了二聚化不良的影响。突变 CGB8 p.Pro73Arg（rs72556345）在五个杂合子（三个 RM 病例和两个对照个体）中发现，并遗传给七个研究的活产儿中的两个。该突变导致约 50%的分泌β亚基获得替代构象，但不影响其生物学活性。对于 CGB8 p.Arg8Trp（rs72556341）取代，应用体外方法未发现完整 hCG 组装发生变化，计算机模拟分析也支持这一结果。总之，累积的数据表明，只有具有中性或轻度功能后果的突变可能在主要的 hCGβ 基因 CGB5 和 CGB8 中被耐受。
