Discovery Pipeline for Genetic Defects in Hypothalamic-pituitary Development Using International Mouse Phenotyping Consortium Mice

摘要 先天性垂体功能减退症（CH）是一种常见的出生缺陷，经常与综合征相关。 中枢神经系统、眼部结构、面部和性腺的异常。最严重的疾病 有中线发育异常，包括前脑无裂畸形，这通常是胚胎致死，或 视隔发育不良不太严重的出生缺陷只会破坏下丘脑或垂体的发育， 激素缺乏，影响生存能力、生长、生育力、代谢和应激反应， 需要终身护理。已知有超过60种基因会导致CH，其中许多是在小鼠中首次发现的。 尽管如此，81%的CH患者仍然缺乏分子诊断，这对计划制定非常重要 治疗和预测未来的风险。我们提出了现有胚胎致死基因敲除的详细表型 已知遗传缺陷导致下丘脑和/或垂体畸形的小鼠。这将扩大 CH和相关的人类中线缺陷的分子诊断，并增加我们的 了解这些关键组织的器官发生。我们确定了一组18个胚胎 下丘脑和/或脑垂体明显畸形的致死或亚存活小鼠系。这 一组基因根据功能进行分组，包括表观遗传调节因子、纤毛成分、蛋白质 修改和其他新的功能类别。我们组建了一个调查小组， 下丘脑-垂体发育、小鼠表型和生物信息学。我们将进行深入和 在选定的IMPC小鼠中对下丘脑和脑垂体进行彻底的表型分析。我们的管道 三个步骤，在每个阶段适当重新确定优先级。第一，时间和空间的表达 选择的基因将在野生型小鼠中确定，IMPC菌株将进口， 从妊娠中期到出生，畸形将以组织学为特征。第二，压力将 评估血管化缺陷和下丘脑神经元和垂体激素的特化 使用scRNA-seq和组织学方法的组合产生细胞。第三，机械化 对发育缺陷的病理生理学的理解将使用组织学、细胞学和免疫组织化学来确定。 培养测定和/或建立的组织特异性cre驱动程序，以测试从scRNA-seq 和荟萃分析。这项工作将建立新基因的基因型/表型关系， 未确诊的CH患者的候选人。数据传播将及时和公开，建立在我们的基础上， 现有的小鼠脑垂体分析数据库（mousePAD）和博客，以及IMPC网站。未来扩展 我们的管道将增加额外的致命或亚活性IMPC菌株，这些菌株的基因突变涉及 下丘脑-垂体发育，但尚未研究。我们准备对世界产生重大影响， 发现影响小鼠和人类下丘脑-垂体发育的遗传缺陷。

Abstract Congenital hypopituitarism (CH) is a common birth defect frequently associated with syndromic abnormalities in the central nervous system, ocular structures, face, and gonads. The most severe disorders have midline developmental anomalies and include holoprosencephaly, which is usually embryonic lethal, or septo-optic dysplasia. Less severe birth defects disrupt only hypothalamic or pituitary development, causing hormone deficiencies that affect viability, growth, fertility, metabolism, and the stress response, and may require life-long care. Over 60 genes are known to cause CH, many of which were first discovered in mice. Nonetheless, 81% of CH patients still lack a molecular diagnosis, which would be invaluable for planning treatment and predicting future risk. We propose detailed phenotyping of existing embryonic lethal knockout mice with known genetic defects that result in hypothalamic and/or pituitary malformations. This will expand the molecular diagnoses for CH and associated midline deficiencies in humans and increase our understanding of organogenesis of these critical tissues. We identified a prioritized set of 18 embryonic lethal or sub-viable mouse lines with obvious malformations in the hypothalamus and/or pituitary gland. This set of genes are grouped based on function including epigenetic regulators, components of cilia, protein modification, and other novel functional categories. We assembled a team of investigators with expertise in hypothalamic-pituitary development, mouse phenotyping, and bioinformatics. We will conduct deep and thorough phenotyping of the hypothalamus and pituitary gland in the selected IMPC mice. Our pipeline has three steps with appropriate re-prioritization at each stage. First, temporal and spatial expression of the selected genes will be determined in wild type mice, the IMPC strains will be imported, and the dysmorphology will be characterized histologically from mid- gestation to birth. Second, the strains will be assessed for defects in vascularization and specification of hypothalamic neurons and pituitary hormone producing cells using a combination of scRNA-seq and histological methods. Third, a mechanistic understanding of the pathophysiology of the developmental defects will be determined using histology, cell culture assays and/or established tissue-specific cre drivers to test hypotheses developed from scRNA-seq and meta-analyses. This work will establish genotype/phenotype relationships for novel genes that are candidates for the undiagnosed CH patients. Data dissemination will be timely and open, building on our existing mouse pituitary analysis database (mousePAD) and blog, and the IMPC website. Future expansion of our pipeline will add additional lethal or sub-viable IMPC strains with mutations in genes implicated in hypothalamic-pituitary development, but not yet studied. We are poised to make a significant impact on the discovery of genetic defects that affect hypothalamic-pituitary development in mouse and man.