Molecular Mechanism of THM1-Medicated Renal Cystogenesis

囊性肾病（CKD）被认为起源于潜在的纤毛缺陷，但分子机制仍不清楚。我们已经确定了一种新的纤毛蛋白，THM 1（Tetratricopeptide Repeat Containing Hedgehog Modulator 1，也称为TTC 21 B或IFT 139），它负调节Hedgehog（Hh）信号。近年来，研究发现，THM 1是纤毛疾病中致病性最强的等位基因，包括Meckel-Gruber综合征（MKS）、Bardet-Bledl综合征（BBS）、肾单位结核（NPHP）、Joubert综合征（JS）和Jeune窒息性胸廓疾病（JATD）。这些纤毛病的主要临床特征是CKD，并且实际上，在晚期胚胎发生期间Thml的遗传缺失导致成年小鼠中的CKD。虽然Hh信号转导在CKD中尚未被广泛研究，但我们对THM 1的分析使我们研究了Hh活性增强在肾囊肿形成中的可能作用。为了支持这一假设，在CKD的胚胎培养模型中，通过遗传缺失Gli 2（Hh信号传导的主要转录激活因子）和用小分子Hh抑制剂处理来防止囊肿形成。本提案的目的是确定是否增强Hh活性导致肾囊肿。在第一个目标中，CKD中旁分泌Hh信号传导的作用将通过在Thm 1条件性敲除小鼠中进行Hh信号传导的空间和时间定量评估来阐明。在第二个目标中，将通过检查在肾小管上皮细胞与肾间质细胞中消融Thm 1的影响来扩展这种空间分析。在第三个目标中，将通过在Thm 1条件性基因敲除小鼠中下调Hh通路的遗传和转录来探索Hh信号传导增加在肾囊肿形成中的因果关系。在空间背景下检查基因表达可能提供一种工具，以更好地了解CKD的发病机制。此外，这些实验将建立一个因果关系的Hh信号在肾脏囊肿的作用，并可能提供重要的影响，对慢性肾脏病的预防策略的设计。

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.