Section on Molecular Endocrinology, Division of Developmental Biology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, USA.
Transgenic Core, National Institute of Mental Health, National Institutes of Health, Bethesda, MD, USA.
Hum Mol Genet. 2019 Aug 1;28(15):2561-2572. doi: 10.1093/hmg/ddz079.
Gonadotropin-regulated testicular RNA helicase (GRTH/DDX25) is a testis specific member of the DEAD-box family of RNA helicases expressed in meiotic and haploid germ cells which plays an essential role in spermatogenesis. There are two species of GRTH the 56 kDa non-phospho and 61 kDa phospho forms. Our early studies revealed a missense mutation (R242H) of GRTH in azoospermic men that when expressed in COS1-cells lack the phospho-form of GRTH. To investigate the role of the phospho-GRTH species in spermatogenesis, we generated a GRTH knock-in (KI) transgenic mice with the R242H mutation. GRTH-KI mice are sterile with reduced testis size, lack sperm with spermatogenic arrest at round spermatid stage and loss of the cytoplasmic phospho-GRTH species. Electron microscopy studies revealed reduction in the size of chromatoid bodies (CB) of round spermatids (RS) and germ cell apoptosis. We observed absence of phospho-GRTH in the CB of RS. Complete loss of chromatin remodeling and related proteins such as TP2, PRM2, TSSK6 and marked reduction of their respective mRNAs and half-lives were observed in GRTH-KI mice. We showed that phospho-GRTH has a role in TP2 translation and revealed its occurrence in a 3' UTR dependent manner. These findings demonstrate the relevance of phospho-GRTH in the structure of the chromatoid body, spermatid development and completion of spermatogenesis and provide an avenue for the development of a male contraceptive.
促性腺激素调节的睾丸 RNA 解旋酶 (GRTH/DDX25) 是 DEAD 盒家族中一种睾丸特异性 RNA 解旋酶，在减数分裂和单倍体生殖细胞中表达，在精子发生中发挥重要作用。GRTH 有两种形式：56kDa 非磷酸化和 61kDa 磷酸化形式。我们的早期研究揭示了 GRTH 在无精子症男性中的一个错义突变（R242H），当在 COS1 细胞中表达时，缺乏 GRTH 的磷酸化形式。为了研究磷酸化-GRTH 形式在精子发生中的作用，我们生成了具有 R242H 突变的 GRTH 敲入 (KI) 转基因小鼠。GRTH-KI 小鼠不育，睾丸体积减小，缺乏精子，精子发生停滞在圆形精子阶段，细胞质磷酸化-GRTH 形式缺失。电子显微镜研究显示，圆形精子 (RS) 的染色质体 (CB) 大小减小，生殖细胞凋亡。我们观察到 RS 的 CB 中缺乏磷酸化-GRTH。在 GRTH-KI 小鼠中，完全缺失染色质重塑和相关蛋白，如 TP2、PRM2、TSSK6，及其各自的 mRNA 和半衰期明显减少。我们表明，磷酸化-GRTH 在 TP2 翻译中发挥作用，并揭示其以 3'UTR 依赖的方式发生。这些发现表明磷酸化-GRTH 在染色质体的结构、精子发生和精子发生完成中具有重要作用，并为开发男性避孕药提供了途径。
