Role of TRIM71 in neural stem cell biology and congenital hydrocephalus

项目概要 先天性脑积水（CH）是最常见的大脑发育畸形，影响 1/1000 的出生率。 CH 通常归因于脑脊液 (CSF) 稳态失常和 因此，通过手术脑脊液改道进行治疗，发病率和失败率很高。后的坚持 许多患者的手术脑室扩大，通常神经发育结果不佳，引发了问题 关于我们当前的 CH 范式及其治疗。我们对分子的理解存在重大差距 CH 的发病机制阻碍了预防措施、靶向治疗和改进的发展 预测。通过全外显子组测序，Kahle 实验室从头鉴定出杂合子 TRIM71/lin-41 突变是人类散发性慢性肝炎最常见的遗传原因（Jin 等，2017）。 2020，自然医学）。 TRIM71 编码一种 RNA 结合蛋白，首次在秀丽隐杆线虫中发现 调节表皮干细胞发育时间的异时基因。尽管其非凡 整个系统发育的进化保守性和产前神经干细胞（NSC）的稳健表达， TRIM71 在哺乳动物大脑发育和 CH 中的作用基本上是未知的。在前期工作中，我 已经表明，具有人类 TRIM71 CH 突变 R608H 的鼠同源物的小鼠品系 (Trim71R595H/ ) 或条件性 NSC 特异性 Trim71 缺失 (Nestin-Trim71fl/fl) 均重述 携带 TRIM71 的人类患者的严重新生儿交通性脑积水 突变。我建议的目标是进一步描述这些模型的特征，以增加对 人类大脑发育和 CH 发病机制。我假设 TRIM71 突变型脑室扩大的结果不是 来自原发性脑脊液（CSF）过度积累，而是与相关的神经发生受损 减少 NSC 增殖和早熟 NSC 分化。目标 1 将描述神经解剖学特征 使用脑磁共振成像 (MRI) 和直接技术研究 TRIM71 突变型脑积水的 CSF 生理学 脑脊液流体动力学的测量。目标 2 将阐明 TRIM71 突变体的细胞机制 使用 Trim71 CH 突变小鼠的免疫荧光研究并辅以体外测定来检测脑积水 使用原代 NSC 培养物。证明神经发生失调而不是原发性脑脊液失调 一些CH案例背后的过度积累可能会产生范式改变的影响，从而可能导致 改进诊断、预后和治疗策略，包括预测哪些 CH 患者 神经外科脑脊液分流术可能会或可能不会受益。这些研究将作为我们的一部分进行 开发CH精准医学疗法的长期目标。

PROJECT SUMMARY Congenital hydrocephalus (CH) is the most common developmental malformation of the brain affecting 1/1000 births. CH has been classically attributed to failed cerebrospinal fluid (CSF) homeostasis and therefore treated by surgical CSF diversion, with high morbidity and failure rates. The persistence of post- surgical ventriculomegaly in many patients, often with poor neurodevelopmental outcomes, raises questions about our current paradigms of CH and its treatment. Significant gaps in our understanding of the molecular pathogenesis of CH impede the development of preventive measures, targeted therapies, and improved prognostication. By whole exome sequencing, the Kahle lab has identified heterozygous de novo mutations in the TRIM71/lin-41 as the most common genetic cause of human sporadic CH (Jin et al. 2020, Nature Medicine). TRIM71 encodes an RNA-binding protein first discovered in C. elegans as a heterochronic gene that regulates developmental timing of epidermal stem cells. Despite its remarkable evolutionary conservation across phylogeny and robust expression in prenatal neural stem cells (NSCs), the roles of TRIM71 in mammalian brain development and CH are essentially unknown. In preliminary work, I have shown that mouse lines with the murine homolog of the human TRIM71 CH mutation R608H (Trim71R595H/+) or with conditional NSC-specific Trim71 deletion (Nestin-Trim71fl/fl) both recapitulate the severe neonatal-onset communicating hydrocephalus of human patients harboring TRIM71 mutations. The objective of my proposal is to further characterize these models to increase knowledge of human brain development and CH pathogenesis. I hypothesize TRIM71 mutant ventriculomegaly results not from primary cerebrospinal fluid (CSF) over-accumulation, but rather from impaired neurogenesis related to reduced NSC proliferation and precocious NSC differentiation. Aim 1 will characterize the neuroanatomy and CSF physiology of TRIM71-mutant hydrocephalus using brain magnetic resonance imaging (MRI) and direct measurements of CSF hydrodynamics. Aim 2 will elucidate the cellular mechanisms of TRIM71-mutant hydrocephalus using immunofluorescent studies in Trim71 CH mutant mice complemented by in vitro assays using primary NSC cultures. The demonstration that dysregulated neurogenesis rather than primary CSF over-accumulation underlies some CH cases could have paradigm-changing implications that could lead to improved diagnostic, prognostic, and therapeutic strategies, including the prediction of which CH patients may or may not benefit from neurosurgical CSF shunting. These studies will be conducted as part of our long-term goal to develop precision medicine therapies for CH.