Biochemical and Functional Studies of Sall1 in Kidney Development and TBS

描述（由申请人提供）：器官发生需要对基因表达进行精确的时间和空间控制，以指导调节各种细胞过程的发育程序。小基因编码多种锌指转录因子，这些因子是器官发育的关键保守调节因子。SALL1基因突变导致汤恩斯-布罗克斯综合征（TBS），这是一种常染色体显性遗传病，伴有影响多个器官的发育缺陷。在这种情况下，肾脏异常是突出的。我们建立了小鼠TBS模型，并表明这种综合征是由Sall1N截断突变蛋白的显性效应引起的。这些小鼠表现出肾发育不全，输尿管分支减少，与TBS患者相似。在Sall1纯合突变小鼠中，输尿管不能正常启动分支，导致肾脏发育不全。虽然这些研究表明Sall1对肾脏发育至关重要，但其功能的分子和发育机制尚不清楚。为了描述Sall1在肾脏中的功能，确定Sall1靶基因并确定这些下游通路如何调节输尿管芽的分支形态发生、后肾祖细胞的维持和肾元的形成是很重要的。我们在Sall1 （SRM）中发现了一个新的抑制基序，该基序是招募核小体重塑和去乙酰化酶（NuRD）复合物介导Sall1靶基因抑制的必要和充分条件。我们目前的模型是，在肾脏发育过程中，Sall1靶基因的一个重要亚群的调控取决于NuRD的募集。根据我们的初步数据，Sall1在肾脏形成过程中可能在多个步骤中起作用。然而，现有的Sall1等位基因所表现出的后肾发育的早期停滞限制了对这些过程的研究。由于Sall1在帽间质（后肾祖细胞）和间质细胞中均有表达，因此确定其在这些不同细胞区室和不同发育阶段的功能是该领域的一个重要问题。这些想法将在三个具体目标中得到验证：(1)后肾间质中Sall1直接靶基因的鉴定和功能分析；(2)研究Sall1在发育肾脏的帽间质和间质细胞中的作用；(3)确定胚胎肾脏中Sall1天然蛋白复合物的组成。这些目标将增加我们对Sall1在肾形成和TBS病因学中的功能的理解。公共卫生相关性：影响肾脏和泌尿道的先天性异常是最常见的严重出生缺陷之一。虽然这些疾病有重要的临床和越来越多的遗传数据，但疾病机制尚未得到很好的理解。所提出的研究将阐明这些遗传性器官缺陷的分子发病机制，这反过来可能导致新的治疗方法。

DESCRIPTION (provided by applicant): Organogenesis requires the precise temporal and spatial control of gene expression to direct developmental programs that regulate a variety of cellular processes. Sall genes encode for multi-zinc finger transcription factors that are critical conserved regulators of organ development. Mutations in SALL1 cause the Townes-Brocks syndrome (TBS), an autosomal dominant disorder with developmental defects affecting multiple organs. Anomalies of the kidney are prominent in this condition. We created a mouse model of TBS and showed that this syndrome results from dominant effects of a Sall1N truncated mutant protein. These mice display renal hypoplasia with reduced branching of the ureter, similar to that seen in TBS patients. In Sall1 homozygous mutant mice the ureter fails to properly initiate branching resulting in renal agenesis. While these studies indicate that Sall1 is essential for kidney development, the molecular and developmental mechanisms of its function are not well understood. To delineate the function of Sall1 in the kidney it is important to identify Sall1 target genes and determine how these downstream pathways regulate branching morphogenesis of the ureteric bud, maintenance of metaneprhic progenitors and nephron formation. We discovered a novel repression motif in Sall1 (SRM) that is necessary and sufficient to recruit the nucleosome remodeling and deacetylase (NuRD) complex to mediate repression of native Sall1 target genes. Our current model is that regulation of an important subset of Sall1 target genes in developing kidney depends on NuRD recruitment. Based on our preliminary data it is likely that Sall1 acts at multiple steps during nephrogenesis. However, the early arrest of metanephric development exhibited by existing Sall1 alleles has limited investigation of these processes. Since Sall1 is expressed in both the cap mesenchyme (metanephric progenitors) and stromal cells, defining its function in these distinct cellular compartments and at different developmental stages is an important question in the field. These ideas will be tested in three specific aims: (1) Identification and functional analysis of direct Sall1 target genes in metanephric mesenchyme (2) Investigate the role of Sall1 in cap mesenchyme and stromal cells in developing kidney, and (3) Determine the composition of Sall1 native protein complexes in embryonic kidney. These aims will increase our understanding of Sall1 function in nephrogenesis and the etiology of TBS. PUBLIC HEALTH RELEVANCE: Congenital anomalies affecting the kidney and urinary tract are among the most common serious birth defects. While there is significant clinical and increasing genetic data on these disorders, the disease mechanisms are not well understood. The proposed studies will elucidate the molecular pathogenesis of these inherited organ defects and this in turn may lead to novel therapeutic approaches.