Molecular Mechanism of THM1-Medicated Renal Cystogenesis

细胞性肾病(CKD)被认为起源于潜在的纤毛缺陷，尽管分子机制尚不清楚。我们已经鉴定出一种新的纤毛蛋白THM1(包含Hedgehog调制器1的四肽重复序列，也称为TTC21B或IFT139)，它负调控Hedgehog(HH)信号。近年来，纤毛基因中THM1基因对纤毛疾病的致病作用最强，包括Meckel-Gruber综合征(MKS)、Bardet-Bledl综合征(BBS)、肾病综合征(NPHP)、Joubert综合征(JS)和Jeune‘s窒息胸廓疾病(JATD)。这些纤毛疾病的一个主要临床特征是CKD，确实，在胚胎发育后期ThM1的基因缺失导致成年小鼠的CKD。虽然HH信号在慢性肾脏病中还没有得到广泛的研究，但我们对THM1的分析已经引导我们研究了HH活性增强在肾脏囊变中的可能作用。为了支持这一假设，CKD胚胎培养模型中的包囊形成可以通过HH信号的主要转录激活因子Gli2的基因缺失和小分子HH抑制剂的治疗来防止。这项建议的目标是确定HH活性增强是否会导致肾囊肿。在第一个目标中，将通过对Thm1条件性基因敲除小鼠的HH信号进行时空定量评估来阐明旁分泌HH信号在CKD中的作用。在第二个目标中，这一空间分析将通过检测在肾小管上皮细胞和肾基质细胞中消融Thm1的效果来扩展。在第三个目标中，将通过在Thm1条件性基因敲除小鼠中从遗传和药理学上下调HH途径，来探索HH信号在肾脏囊变中增加的因果关系。在空间背景下检测基因表达可能为更好地了解CKD的发病机制提供了一种工具。此外，这些实验将确定HH信号在肾脏囊变中的因果作用，并可能为设计预防CKD的策略提供重要的启示。

Cysfic kidney disease (CKD) is proposed to originate from an underlying ciliary defect, though molecular mechanisms remain unclear. We have identified a novel ciliary protein, THM1 (Tetratricopeptide Repeat Containing Hedgehog Modulator 1, also termed TTC21B or IFT139), which negatively regulates Hedgehog (Hh) signaling. Recently, THM1 has been shown to contribute the most pathogenic alleles among ciliary genes to patients with ciliopathies, including Meckel-Gruber Syndrome (MKS), Bardet-Bledl Syndrome (BBS), nephronophthisis (NPHP), Joubert's Syndrome (JS) and Jeune's Asphyxiating Thoracic Disorder (JATD). A major clinical feature of these ciliopathies is CKD, and indeed, genetic deletion of Thml during late embryogenesis results in CKD in the adult mouse. While Hh signaling has not been studied extensively in CKD, our analyses of THM1 have led us to investigate a possible role for enhanced Hh activity in renal cystogenesis. In support of this hypothesis, cyst formation in an embryonic culture model of CKD was prevented by genetic deletion of Gli2, the main transcriptional activator of Hh signaling, and by treatment with small molecule Hh inhibitors. The goal of this proposal is to determine whether enhanced Hh activity leads to renal cysts. In the first aim, the effect of paracrine Hh signaling in CKD will be elucidated by conducting a spatial and temporal quantitative assessment of Hh signaling in the Thm1 conditional knock-out mouse. In the second aim, this spatial analysis will be extended by examining the effects of ablating Thm1 in renal tubular epithelial cells versus in renal stromal cells. In the third aim, causality of increased Hh signaling in renal cystogenesis will be explored by down regulating the Hh pathway genetically and pharmacologically in Thm1 conditional knock-out mice. Examining gene expression in a spatial context may provide a tool with which to gain better understanding of the pathogenesis of CKD. Furthermore, these experiments will establish a causal role for Hh signaling in renal cystogenesis and may offer important implications for the design of preventive strategies against CKD.