Detection of <scp>REST</scp> expression in the testis using epitope‐tag knock‐in mice generated by genome editing
Detection of <scp>REST</scp> expression in the testis using epitope‐tag knock‐in mice generated by genome editing
复制标题
使用基因组编辑生成的表位标签敲入小鼠检测睾丸中的 <scp>REST</scp> 表达
DOI：
10.1002/dvdy.417
复制
发表时间：
2021
影响因子：
2.5
通讯作者：
Osumi Noriko
中科院分区：
文献类型：
--
作者：
Kimura Ryuichi;U. Inoue Yukiko;Kikkawa Takako;Tatehana Misako;Morimoto Yuki;Inada Hitoshi;Oki Shinya;Inoue Takayoshi;Osumi Noriko
背景阻遏元件1-沉默转录因子（Repressorelement 1-silencingtranscription factor，REST）是一种在多能干细胞中高度表达的主调节因子，可抑制涉及广泛生物学过程的基因网络。最近的一项研究表明，REST可能参与了老年父亲后代胚胎大脑中靶基因的错误调节。然而，由于REST蛋白在特定细胞类型中的检测困难，因此缺乏对REST在精子发生中的功能的详细分析。结果为了确定REST的定位，我们通过CRISPR/Cas9系统在内源性Restlocus处产生了具有podoplanin（PA）标签的C末端插入的表位标签敲入（KI）小鼠系。PA标签的定位在胚胎脑中标记有Pax 6的神经干细胞中得到证实。此外，PA标记的REST中检测到未分化和分化的精原细胞以及支持细胞在新生儿和成人testes.ConclusionsWe表明，REST的表达在精子发生的早期步骤，并建议一种可能性，REST可能会调节男性生殖细胞的表观遗传状态。我们的KI小鼠可能有助于研究神经发育和年龄相关疾病的REST相关分子机制。
BackgroundRepressor element 1‐silencing transcription factor (REST) is a master regulator that is highly expressed in multipotent stem cells to repress gene networks involving a wide range of biological processes. A recent study has suggested that REST might be involved in a misregulation of its target genes in the embryonic brain of offspring derived from aged fathers. However, detailed analyses of the REST function in spermatogenesis are lacking due to difficulty in the detection of REST protein in specific cell types.ResultsTo determine localization of REST, we generated an epitope tag knock‐in (KI) mouse line with the C‐terminus insertion of a podoplanin (PA)‐tag at an endogenousRestlocus by the CRISPR/Cas9 system. Localization of the PA‐tag was confirmed in neural stem cells marked with Pax6 in the embryonic brain. Moreover, PA‐tagged REST was detected in undifferentiated and differentiating spermatogonia as well as Sertoli cells in both neonatal and adult testes.ConclusionsWe demonstrate that REST is expressed at the early step of spermatogenesis and suggest a possibility that REST may modulate the epigenetic state of male germline cells. Our KI mice may be useful for studying REST‐associated molecular mechanisms of neurodevelopmental and age‐related disorders.