Decoding reno-protective mechanisms in mouse Pkhd1 models: Implications for ARPKD therapeutics

项目总结 在美国约有1500名常染色体隐性遗传性多囊肾病(ARPKD； MIM 173900)，一种肝肾纤维囊性疾病，其特征是肾脏增大，有无数的集合体 导管囊肿和进行性肾功能丧失。基本上，所有的ARPKD病例都可以归因于突变 在PKHD1中，它编码一种单程跨膜蛋白，即纤维囊蛋白/多聚管蛋白(FPC)。我们目前有 对人类ARPKD的发病机制了解甚少，因此，治疗在很大程度上是支持性的。 PKHD1/Pkhd1肾脏表型在物种间的显著差异可能为 疾病机制。而PKHD1错义突变或截短突变的人类患者则有严重的 肾囊性疾病，带有工程截断突变的小鼠Pkhd1模型(以及可能的FPC丢失 功能)表示轻微或无肾脏疾病。我们的初步研究表明，虽然MYC过度表达 在人ARPKD肾脏中，Myc在小鼠Pkhd1肾脏中没有过表达。在之前的研究中，我们有 结果表明，FPC经历了缺口样的加工，从 质膜和核运输。在这里，我们演示了小鼠FPC-CTD直接与 MYC P1启动子，增加Myc的表达。 基于这些发现，我们假设小鼠肾上皮细胞可以补偿fpc-ctd的丢失。 通过肾脏保护机制发挥作用以及物种特异性FPC-CTD对Myc表达的调节 是这种雷诺保护的核心。我们推测，虽然这些机制在人类体内并不正常运行 对于肾上皮细胞，他们可能会发现新的靶向治疗人类ARPKD肾脏疾病的机会。我们 提出两个具体目标来验证我们的假设：(1)确定FPC-CTD如何调节Myc/MYC的表达 在小鼠和人的肾上皮细胞中，并确定所提出的调控电路的中断是否是一种 (2)比较小鼠和人的FPC-CTD核相互作用组学。 和检测转录靶点的差异是否识别小鼠肾脏保护途径 肾脏。 我们的研究将通过顺序解决FPC-CTD的转录作用来推动该领域的发展。具体来说， 我们将：1)确定 小鼠Pkhd1模型； FPC-CTD相关Myc转录调控在肾脏保护中的作用 以及2)确定允许小鼠肾上皮细胞补偿的可能机制 FPC-CTD核功能丧失。展望未来，这些数据将为MICE的翻译奠定基础 将肾脏保护机制转化为减轻人PKHD1相关肾囊肿的新的治疗策略 疾病。

PROJECT SUMMARY There are ~1,500 patients in the United States with autosomal recessive polycystic kidney disease (ARPKD; MIM 173900), a hepatorenal fibrocystic disorder characterized by enlarged kidneys with innumerable collecting duct cysts and progressive loss of renal function. Essentially all cases of ARPKD can be attributed to mutations in PKHD1, which encodes a single-pass transmembrane protein, fibrocystin/polyductin (FPC). We currently have very little insight into the pathogenesis of human ARPKD and thus, treatment is largely supportive. Striking species-specific differences in the PKHD1/Pkhd1 renal phenotype may offer important insights into disease mechanisms. While human patients with either missense or truncating PKHD1 mutations have severe renal cystic disease, mouse Pkhd1 models with engineered truncating mutations (and presumably loss of FPC function) express minimal or no renal disease. Our preliminary studies reveal that while MYC is overexpressed in human ARPKD kidneys, Myc is not overexpressed in mouse Pkhd1 kidneys. In previous studies, we have shown that FPC undergoes Notch-like processing with cleavage of the carboxy terminus (FPC-CTD) from the plasma membrane and nuclear trafficking. Here, we demonstrate that the mouse FPC-CTD binds directly to the Myc P1 promoter and increases Myc expression. Based on these findings, we hypothesize that mouse renal epithelia can compensate for the loss of FPC-CTD function through reno-protective mechanisms and that species-specific, FPC-CTD regulation of Myc expression is central to this reno-protection. We speculate that while these mechanisms are not normally operative in human renal epithelia, they may identify new opportunities for therapeutic targeting in human ARPKD renal disease. We propose two specific aims to test our hypothesis: (1) define how the FPC-CTD regulates Myc/MYC expression in mouse and human renal epithelia and determine whether disruption of the proposed regulatory circuits is a central driver of renal cystogenesis; and (2) compare the FPC-CTD nuclear interactome in mouse and human renal epithelia and test whether differences in transcriptional targets identifies reno-protective pathways in mouse kidneys. Our studies will advance the field by sequentially addressing the transcriptional role of FPC-CTD. Specifically, we will: 1) determine mouse Pkhd1 models; how FPC-CTD related Myc transcriptional regulation contributes to reno-protection in and 2) identify putative mechanisms that allow mouse renal epithelia to compensate for the loss of FPC-CTD nuclear function. Moving forward, these data will lay the foundation for translating mouse reno-protective mechanisms into novel, therapeutic strategies that attenuate human PKHD1-related renal cystic disease.