Transcription Factors in Early Kidney Development

描述（由申请人提供）：本提案的长期目标是通过研究转录辅激活因子Eya1、其相互作用的同源结构域蛋白Six1及其辅因子的作用，阐明在早期肾脏发育过程中控制功能性后肾间质形成的分子机制。哺乳动物肾脏通过后肾间质和输尿管芽上皮之间的诱导相互作用，在后中胚层的一个区域发育。最近的遗传和分子研究表明，后肾间质可能提供促进输尿管芽形成的初始信号。然而，控制后肾间质形成的基因和调控层次尚不清楚。最近，小鼠Eya1基因被发现在后肾间质中表达，而人类Eya1基因的突变可引起鳃裂-耳-肾综合征（BOR），这是一种以鳃裂、耳部和肾脏联合异常为特征的先天性出生缺陷。然而，尽管确定了相关基因，但肾脏缺陷的发育和分子基础以及Eya1在早期肾脏形态发生中的功能步骤尚不清楚。在Eya1 -/-小鼠中，从E10.5开始后肾间质不形成，间质细胞发生异常凋亡，表明Eya1是早期肾脏形态发生所必需的关键间质基因。同源盒基因Six1也被发现在早期肾脏发育中发挥重要作用，其基因产物与Eya1物理相互作用。此外，最近以Eya1为“诱饵”，通过酵母双杂交筛选分离到转录因子N-myc，并在体外与Six1和Eya1发生物理相互作用。有趣的是，N-myc基因的突变也会导致肾脏缺陷。基于这些数据，我们假设这些转录因子在后肾间质中共同起调节早期肾脏发育的作用。这项资助将使用一个强大的遗传系统来测试这一假设，并将几个间充质基因整合到控制早期肾脏发育的遗传和分子调控途径中。具体而言，我建议：(1)测试Eya1和Six1在早期肾脏发育中的功能作用；(2)测试Six1、Eya1和Pax2在早期肾脏发育中可能的相互作用；(3)测试N-myc在早期肾脏发育中是否与Eya1或Six1相互作用。这些研究将阐明Pax2、Eya1、Six1、N-myc等基因之间的关系，并极有可能在分子和遗传水平上对肾脏形态发生的早期发育过程提供有意义的认识。

DESCRIPTION (provided by applicant): The long-term objective of this proposal is to elucidate the molecular mechanisms controlling the formation of a functional metanephric mesenchyme during early kidney development by examining the role of the transcriptional coactivator Eya1, its interacting homeodomain protein Six1 and their cofactors. The mammalian kidney develops in a region of posterior intermediate mesoderm by inductive interactions between the metanephric mesenchyme and the ureteric bud epithelium. Recent genetic and molecular studies have indicated that the metanephric mesenchyme may provide initial signals to promote ureteric bud formation. However, what genes and the regulatory hierarchy controlling the formation of the metanephric mesenchyme still remain unclear. Recently, the murine Eya1 gene was found to be expressed in the metanephric mesenchyme and mutations in the human EYA1 gene cause Branchio-Oto-Renal (BOR) syndrome, a congenital birth defect characterized by combinations of branchial, otic and renal anomalies. However, despite the identification of the responsible gene, the developmental and molecular basis for renal defects and the identity of the steps at which Eya1 functions in early kidney morphogenesis are unclear. In Eya1 -/- mice, the metanephric mesenchyme never forms and the mesenchymal cells undergo abnormal apoptosis from E10.5, indicating that Eya1 is a key mesenchymal gene required for early kidney morphogenesis. The homeobox gene Six1 was also found to play an essential role during early kidney development and its gene product physically interacts with Eya1. Moreover, the transcription factor N-myc has been recently isolated through yeast two-hybrid screen using Eya1 as "bait" and it physically interacts with both Six1 and Eya1 in vitro. Interestingly, mutations in the N-myc gene also cause kidney defects. Based on these data, we hypothesize that these transcription factors function together in the metanephric mesenchyme to regulate early kidney development. This grant will use a powerful genetic system to test this hypothesis and integrate several mesenchymal genes into a genetic and molecular regulatory pathway governing early kidney development. Specifically, I propose to: (1) test the functional role of Eya1 and Six1 during early kidney development, (2) test the possible interactions between Six1, Eya1 and Pax2 in early kidney development, (3) test whether N-myc interacts with Eya1 or Six1 during early kidney development. These studies will clarify the relationship between Pax2, Eya1, Six1, N-myc and other genes, and have a strong likelihood of providing significant insight at the molecular and genetic level into the early developmental process of kidney morphogenesis.