Two ovarian candidate enhancers, identified by time series enhancer RNA analyses, harbor rare genetic variations identified in ovarian insufficiency
Two ovarian candidate enhancers, identified by time series enhancer RNA analyses, harbor rare genetic variations identified in ovarian insufficiency
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通过时间序列增强子 RNA 分析鉴定出两个卵巢候选增强子，它们含有在卵巢功能不全中发现的罕见遗传变异
DOI：
10.1093/hmg/ddac023
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发表时间：
2022
影响因子：
3.5
通讯作者：
Nakagawa Ryuichi et al.
中科院分区：
文献类型：
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作者：
Hayasaka Kazuki;Watanabe Yui;Hirama Takashi;Oishi Hisashi;Noda Masafumi;Toyama Hiroaki;Ejima Yutaka;Saiki Yoshikatsu;Okada Yoshinori;Nakagawa Ryuichi et al.
卵巢发育的基因调控在很大程度上仍不清楚。事实上，在大多数卵巢发育受损的病例中，例如46，XX性发育障碍(DSD)没有SRY，以及卵巢功能不全(POI)-遗传原因尚未确定，绝大多数与疾病相关的序列变异可能位于非编码调控序列中。在这项研究中，我们的目标是确定五个已知在卵巢早期发育中发挥关键作用的卵巢基因的增强子，我们的分析基于增强子衍生转录本(ERNAs)的表达，这些转录被认为是活性增强子的特征。通过对E13.5、E16.5和P0的基因表达进行CAP分析，获得了小鼠WT1阳性卵巢细胞表达谱的时间变化。我们比较了卵巢特异的Erna的时序表达谱和每个卵巢特异基因的表达谱，得到了Wnt4和Rspo1的两个候选序列。这两个序列在小鼠和人之间都是保守的，我们通过在小鼠颗粒细胞中的瞬时表达分析证实了它们的增强子活性。此外，通过对24例卵巢发育受损患者(POI、性腺发育不良和46，XX DSD)的该区域进行测序，我们在这两个序列中都发现了罕见的单核苷酸变异。我们的结果表明，联合分析靶基因的Erna和mRNA的时间表达谱为定位顺式元件增强子提供了一个强大的工具，并为识别位于非编码调控序列中的疾病相关序列变体提供了一种手段，从而促进了人类遗传学向前发展中一个重要的未得到满足的需求。
The genetic regulation of ovarian development remains largely unclear. Indeed, in most cases of impaired ovarian development—such as 46,XX disorders of sex development (DSD) withoutSRY, and premature ovarian insufficiency (POI)—the genetic causes have not been identified, and the vast majority of disease-associated sequence variants could lie within non-coding regulatory sequences. In this study, we aimed to identify enhancers of five ovarian genes known to play key roles in early ovarian development, basing our analysis on the expression of enhancer derived transcripts (eRNAs), which are considered to characterize active enhancers. Temporal expression profile changes in mouse WT1-positive ovarian cells were obtained from cap analysis of gene expression at E13.5, E16.5 and P0. We compared the chronological expression profiles of ovarian-specific eRNA with expression profiles for each of the ovarian-specific genes, yielding two candidate sequences for enhancers ofWnt4andRspo1.Both sequences are conserved between mouse and human, and we confirmed their enhancer activities using transient expression assays in murine granulosa cells. Furthermore, by sequencing the region in patients with impaired ovarian development in 24 patients, such as POI, gonadal dysgenesis and 46,XX DSD, we identified rare single nucleotide variants in both sequences. Our results demonstrate that combined analysis of the temporal expression profiles of eRNA and mRNA of target genes presents a powerful tool for locatingcis-element enhancers, and a means of identifying disease-associated sequence variants that lie within non-coding regulatory sequences, thus advancing an important unmet need in forward human genetics.