The role of DNA damage response signaling in chronic kidney disease

摘要 DNA损伤反应信号在慢性肾脏疾病中的作用。肾单位增多症(NPHP) 一种常染色体隐性遗传性慢性肾病(CKD)，以肾小管间质纤维化为特征 基底膜破裂和肾囊肿。肾纤维化是终末期的主要决定因素 肾脏疾病，目前还没有有效的治疗方法。最近的研究发现，NPHP是一种 “纤毛病”，由基因突变引起，其蛋白质定位于中心体和/或主纤毛， 并产生了NPHP相关纤毛疾病(NPHP-RC)的统一病原学概念。尽管 确认30多个基因突变是NPHP-RC的原因，对该分子知之甚少 这种疾病的潜在机制。最近，通过对整个外显子重新测序，我参与了 3个新基因CEP164、FAN1和SDCCAG8的鉴定与肾性NPHP-RC相关 突变时的变性/纤维化。令人惊讶的是，中心体蛋白CEP164和SDCCAG8也是 定位于细胞核，其失活导致DNA损伤反应(DDR)信号受损 伴有细胞周期缺陷。因此，与已知的DNA损伤修复蛋白FAN1一起， CEP164和SDCCAG8提示DDR信号缺陷是NPHP-RC的一种新的发病机制。 我假设NPHP-RC中的肾脏变性/纤维化是由DNA损伤反应缺陷引起的 小路。为了验证这一假设，我将(1)研究“纤毛病变蛋白”在调节DNA中的作用。 使用Cep164和Sdccag8小鼠肾纤维化模型的损伤反应途径；(2)执行 影响和抵消NPHP-RC的小分子的高通量筛查 (3)CEP164和SDCCAG8的功能和转运调控 它们在中心体和细胞核的双重定位之间的差异。拟议研究的完成情况 将为(1)纤毛疾病的发病机制提供新的见解；(2)能够 导致NPHP-RC和肾脏纤维化的治疗；(3)研究DDR相关疾病的新动物模型。 此K99/R00应用程序的目的是利用新生成的动物模型来研究 NPHP-RC的分子机制。这将有助于我的研究生涯向 独立调查员职位。该奖项的培训(K99)阶段将由Friedhelm博士指导 希尔德布兰特是霍华德·休斯医学研究所的研究员，也是国际公认的 人类遗传学和儿童肾脏疾病领域。2013年3月，我将搬到波士顿。 儿童医院(BCH)/哈佛医学院(HMS)与希尔德布兰特博士的实验室。我的长期生活 职业目标是将自己确立为肾脏研究领域的独立研究员，专注于 肾脏疾病的动物模型。BCH/HMS有几个合作研究小组致力于 肾脏发育和疾病为我的培训和职业发展提供了理想的环境。

ABSTRACT The role of DNA damage response signaling in chronic kidney disease. Nephronophthisis (NPHP) is an autosomal recessive chronic kidney disease (CKD), characterized by tubulointerstitial fibrosis, tubular basement membrane disruption and kidney cysts. Renal fibrosis is the primary determinant of end-stage kidney disease, with no effective therapy available today. Recent work has identified NPHP as a “ciliopathy”, caused by mutations in genes, whose proteins localize to the centrosome and/or primary cilium, and has generated a unifying pathogenic concept for NPHP-related ciliopathies (NPHP-RC). Despite the identification of mutations in more than 30 genes as causative of NPHP-RC, little is known about the molecular mechanisms underlying the disease. Recently, by whole exome resequencing I was involved in the identification of 3 novel genes – CEP164, FAN1 and SDCCAG8 as causing NPHP-RC with renal degeneration/fibrosis when mutated. Surprisingly, the centrosomal proteins CEP164 and SDCCAG8 were also localized at cell nuclei, and their inactivation led to impaired DNA damage response (DDR) signaling accompanied by cell cycle defects. Hence, together with FAN1, which is a known DNA damage repair protein, CEP164 and SDCCAG8 implicate defective DDR signaling as a novel pathomechanism of NPHP-RC. I hypothesize that renal degeneration/fibrosis in NPHP-RC is caused by defects in DNA damage response pathways. To test this hypothesis, I will (1) examine the role of “ciliopathy proteins” in regulating the DNA damage response pathway using the Cep164 and Sdccag8 mouse models of renal fibrosis; (2) perform a high-throughput screen for small molecules that influence and counteract the NPHP-RC pathogenesis; (3) characterize the functions and regulation of trafficking of CEP164 and SDCCAG8 between their dual localizations at centrosomes and in nuclei. Accomplishment of the proposed research will provide (1) new insights into the pathomechanisms of ciliopathies; (2) new chemical compounds that can lead to therapies for NPHP-RC and renal fibrosis; (3) new animal models for studying DDR-related diseases. The objective of this K99/R00 application is to utilize the newly generated animal models for the study of molecular mechanisms of NPHP-RC. This will facilitate the transition of my research career towards an independent investigator position. The training (K99) phase of this award will be mentored by Dr. Friedhelm Hildebrandt, who is an investigator of Howard Hughes Medical Institute and internationally recognized leader in the fields of human genetics and of pediatric kidney diseases. In March 2013 I will move to the Boston Children's Hospital (BCH)/Harvard Medical School (HMS) with Dr. Hildebrandt's laboratory. My long-term career goal is to establish myself as an independent investigator in the field of kidney research, focusing on animal models of kidney diseases. The BCH/HMS has several collaborative research groups working on kidney development and diseases providing an ideal environment for my training and career development.