Integrative Approaches to Decipher Genetic Determinants of Disease Penetrance in Prokineticin 2 Pathway Related Human Reproductive Disorders

实现基因组精确医学的一个主要挑战是观察到基因并不 总是预测表型。不完全外显和可变表现力就是这方面的两个例子 表型变异性。这种复杂性在人类PROK2(编码原激动素)的突变中得到了很好的说明 2)和PROKR2(原激动素受体2)，引起人分离的GnRH(促性腺激素释放激素) 缺乏(IGD)。原激动素2信号是哺乳动物生殖的主要调节因子，并与 人类有神经发育和神经内分泌类型的IGD(Kallmann综合征和正常人 分别为IGD)。除了这种生理上的复杂性，PROK2/R2突变主要是错误的 表现出外显性降低和/或表达能力可变的突变。这种变异性的遗传基础 目前尚不完全清楚，但已推测等位基因异质性和修饰基因可解释这一现象。 可变性。在特定目标1中，将使用以下方法在体外检测人类PROKR2突变的等位基因异质性 PROK2信号分析和等位基因特异性表达研究。全外显子组测序和SNP基因分型 将确定影响疾病外显性和表现力的修饰基因。PROK2还牵涉到 不同的生理过程，如葡萄糖代谢、能量平衡、食物摄入、昼夜节律、疼痛 调节、中性粒细胞趋化和血管生成。这些多效性功能与人类的相关性 PROK2尚未建立。在特定目标2中，对带有PROK2/R2突变的人进行表型解剖 将确定与人类PROK2/R2突变相关的多效性表型。IGD患者 此外，还将研究三级医院生物库种群中的对照个体。生殖表型 而非生殖表型将被绘制成图表。有针对性的表型鉴定将包括葡萄糖的评估 两组均有动态平衡和疼痛调节作用。将在全球范围内进行一项关联研究 含有PROK2/R2突变的生物库受试者。在特定目标3中，使用一种新的PROKR2-cre小鼠模型， PROKR2和GnRH神经元的转录研究将在不同的发育时间点进行，使用 使用化学遗传学对PROKR2的时间抑制。这些小鼠性成熟和生育力在颞叶的变化 还将检查PROKR2抑制情况。在小鼠体内差异表达的顶级基因 转录实验将通过评估这些基因是否出现在 修饰基因在人类整个外显子组测序研究中的应用。这项建议中的研究具有潜在的 揭开与PROK2途径突变相关的疾病外显的新遗传机制 与该途径突变相关的潜在多向性人类表型有助于提供个性化的 为患有生殖障碍和不孕不育的人类提供药物。

A major challenge in accomplishing genomic precision medicine is the observation that genotype does not always predict phenotype. Incomplete penetrance and variable expressivity represent two examples of this phenotypic variability. This complexity is well-illustrated by human mutations in PROK2 (encoding prokineticin 2) and PROKR2 (prokineticin receptor 2), that cause human Isolated GnRH (gonadotropin-releasing hormone) Deficiency (IGD). Prokineticin 2 signaling is a master regulator of mammalian reproduction and is associated with neurodevelopmental and neuroendocrine forms of IGD in humans (Kallmann syndrome, and normosmic IGD, respectively). In addition to this physiologic complexity, PROK2/R2 mutations are predominantly missense mutations that exhibit either reduced penetrance and/or variable expressivity. The genetic basis of this variability is not fully understood but allelic heterogeneity and modifier genes have been postulated to account for this variability. In Specific Aim 1, allelic heterogeneity of human PROKR2 mutations will be examined in vitro, using PROK2 signaling assays and allele specific expression studies. Whole exome sequencing and SNP genotyping will identify modifier genes that contribute to disease penetrance and expressivity. PROK2 is also implicated in diverse physiologic processes such as glucose metabolism, energy balance, food intake, circadian rhythms, pain modulation, neutrophil chemotaxis and angiogenesis. The human relevance of these pleotropic functions of PROK2 is yet to be established. In Specific Aim 2, phenomic dissection of humans with PROK2/R2 mutations will be undertaken to define the pleotropic phenotypes relating to PROK2/R2 mutations in humans. IGD patients as well as control individuals in a tertiary hospital biobank population will be studied. Reproductive phenotypes and non-reproductive phenotypes will be charted. Targeted phenotyping will include assessment of glucose homeostasis and pain modulation in both groups. A phenome-wide association study will be conducted in the biobank subjects harboring PROK2/R2 mutations. In Specific Aim 3, using a novel PROKR2-cre mouse model, transcriptomic studies of PROKR2 and GnRH neurons will be done at various developmental timepoints using temporal inhibition of PROKR2 using chemogenetics. Sexual maturation and fertility of these mice after temporal PROKR2 inhibition will also be examined. The top genes that are differentially expressed in the murine transcriptomic experiments will be translationally validated by assessing whether these genes emerge as modifier genes in the human whole exome sequencing studies. The studies in this proposal have the potential to unravel novel genetic mechanisms of disease penetrance relating to the PROK2 pathway mutations, uncover potential pleotropic human phenotypes associated with mutations in this pathway and help deliver personalized medicine to humans with reproductive disorders and infertility.