Role of GLIS2 in the Development of Cystic Kidney Disease

描述(申请人提供)：肾单位肥厚症(NPHP)是一组常染色体，是导致慢性肾功能衰竭的最常见的遗传原因之一。 以进行性肾囊性变性和纤维化为特征的隐性疾病。GLIS2/NPHP7基因突变是人类和小鼠7型肾炎的原因。GLIS2是一个与果蝇的脊椎动物同源基因GLI1、GLI2和GLI3高度同源的转录因子。与果蝇Ci一样，Gli蛋白是脊椎动物Hedgehog(HH)信号通路的关键分子，HH信号通路在进化中高度保守，在胚胎和出生后生命的增殖、分化和修复调节中发挥核心作用。我们最近已经验证并证实了Glis2是出生后肾脏中HH信号的抑制因子的假说，并证明该通路的功能障碍会导致肾囊肿和纤维化，炎性细胞浸润和细胞凋亡增加，但导致这种肾脏表型的分子事件仍不清楚。在我的实验室进行的初步研究意外地发现，Glis2的缺失导致Toll样受体2(Toll like Receptor 2，TLR2)和促炎症TLR/NF-？B途径的其他成分在肾上皮细胞中过度表达，并导致该途径依赖于TLR2的激活。我们还发现一些miRNAs在肾上皮细胞中的表达受Glis2控制，这表明microRNAs是小鼠肾脏中Glis2/HH信号的效应者。我们建议通过产生TLR2-/-；Glis2mut/mut和Myd88flx/Flox；Glis2mut/mut；KspCre双突变体，并建立TLR2-/-；Glis2mut/mut骨髓嵌合体，在体内测试抑制TLR-2/NF-β信号转导对Glis2mut/mut小鼠肾脏炎症浸润和纤维化的影响。此外，我们建议通过染色质免疫沉淀-高度平行测序(ChIP-Seq)和高通量microRNA测序(miRNA-Seq)在全基因组水平上系统地鉴定规范和非规范的Glis2靶基因。根据我们的最新结果，识别在Glis2基因敲除肾脏中解除调控的下游效应器变得更加重要，并可能对我们的囊性肾脏疾病和纤维化模型中改变的复杂调控网络产生进一步的见解。

DESCRIPTION (provided by applicant): Nephronophthisis (NPHP), one of the most frequent genetic causes of chronic renal failure in the first three decades of life, is a group of autosomal recessive diseases characterized by progressive kidney cystic degeneration and fibrosis. Mutations in the gene GLIS2/NPHP7 are the cause of nephronophthisis type 7 both in humans and mice. GLIS2 is a transcription factor with high sequence homology to GLI1, GLI2 and GLI3, the vertebrate orthologs of Drosophila Ci (cubitus interruptus). Like Drosophila Ci, GLI proteins are key molecules in the vertebrate Hedgehog (Hh) signaling, a pathway highly conserved in the evolution and central in the regulation of proliferation, differentiation and repair during embryonc and postnatal life. We have recently tested and confirmed the hypothesis that Glis2 is a repressor of Hh signaling in the postnatal kidney and demonstrated that malfunctioning of this pathway results in kidney cysts and fibrosis, increased inflammatory infiltration and apoptosis, but the molecular events leading to this kidney phenotype are still unexplained. Preliminary studies performed in my laboratory have unexpectedly revealed that loss of Glis2 results in over expression of Toll like receptor 2 (Tlr2) and other components of the pro-inflammatory TLR/NF- ?B pathway in kidney epithelial cells and in the Tlr2-dependent activation of this pathway. We have also found that expression of several miRNAs in kidney epithelial cells is controlled by Glis2, suggesting that microRNAs are effectors of Glis2/Hh signaling in mouse kidneys. We propose to test in vivo the effect of the inhibition of TLR-2/NF-?B signaling on the inflammatory infiltration and fibrosis in Glis2mut/mut mice kidneys by generating Tlr2-/-;Glis2mut/mut and Myd88flox/flox;Glis2mut/mut;KspCre double mutants and establishing Tlr2-/-;Glis2mut/mut bone marrow chimeras. In addition, we propose to systematically identify at genome wide level the canonical and non-canonical Glis2 target genes by chromatin immunoprecipitation-highly parallel sequencing (ChIP-Seq) and high-throughput microRNA sequencing (miRNA-Seq). In light of our most recent results, identifying downstream effectors that are deregulated in Glis2 knockout kidneys has gained even more importance, and is likely to produce further insights in the complex regulatory network that is altered in our model of cystic kidney diseases and fibrosis.