Mechanism of Hedgehog signal transduction

描述（由申请人提供）：Hedgehog （Hh）家族成员介导局部细胞-细胞通信，这对许多组织和结构的发育和维持至关重要。Hh信号的异常激活是基底细胞癌、成神经管细胞瘤的基础，并可能导致许多其他癌症。Hh信号的转导尚不清楚。Smoothened （Smo）是蛇形受体家族的一员，对于激活细胞内对Hh的反应至关重要。反应通过胞质复合体进行转导，包括Costal（发散激酶）、Fused （S/T激酶）和Ci（锌指转录因子）。根据Hh输入的水平，复合物可以将Ci以潜伏形式（Ci155）保持在细胞质中，允许Ci155进入细胞核，促进Ci的转录激活，或促进Ci的蛋白水解加工成其转录抑制物形式（CiR）。我们最近发现Smo结合细胞质调节复合体来调节其活性。这支持了我们的基因分析模型。该模型表明Smo可以根据Hh水平采取三种不同的状态（OFF， LOW， HIGH）， Smo直接接触Costal和Fused， Smo二聚体是HIGH信号传导所必需的。本实验通过以下方法验证并扩展了这一模型：1)表征在OFF、LOW和HIGH状态下的Ci调控复合体；2)确定Smo二聚化/寡聚化在信号传导中的作用；3)利用内源性Smo对信号传导起主导作用的多种Smo突变形式来识别并进一步剖析信号转导的步骤。这些实验研究了三种不同的信号状态对不同水平Hh的反应的结构和机制基础。他们应该确定Hh信号转导通路三种状态之间转换的关键步骤。他们的成功完成将极大地促进我们对这一基本信号转导途径的理解，并可能确定基于hh的病理的治疗靶点。

DESCRIPTION (provided by applicant): Signaling by Hedgehog (Hh) family members mediate local cell-cell communication that is essential for development and maintenance of many tissues and structures. Aberrant activation of Hh signaling underlies basal cell carcinoma, medulloblastoma, and may contribute to many other cancers. Transduction of the Hh signal is poorly understood. Smoothened (Smo), a member of the serpentine receptor family, is essential for activation of intracellular responses to Hh. Responses are transduced through a cyoplasmic complex including Costal (divergent kinesin), Fused (S/T kinase), and Ci (zinc finger transcription factor). Depending on the level of Hh input, the complex can hold Ci in the cytoplasm in a latent form (Ci155), allow Ci155 to enter the nucleus, promote transcriptional activation by Ci, or promote proteolytic processing of Ci to its transcriptional repressor form (CiR). We have recently shown that Smo binds the cytoplasmic regulatory complex to regulate its activity. This supports our model, derived from genetic analysis. The model suggests that Smo can adopt three distinct states depending on Hh levels (OFF, LOW, HIGH), that Smo directly contacts Costal and Fused, and that Smo dimers are necessary for HIGH signaling. The proposed experiments test and extend this model by 1) characterizing the Ci regulatory complex in the OFF, LOW, and HIGH states, 2) determining the role of Smo dimerization/oligomerization in signaling, and 3) using a variety of mutated forms of Smo with dominant effects on signaling by endogenous Smo to identify and further dissect the steps in signal transduction. These experiments investigate the structural and mechanistic basis for three distinct signaling states in response to different levels of Hh. They should identify the critical steps for transition between the three states of the Hh signal transduction pathway. Their successful completion would significantly advance our understanding of this fundamental signal transduction pathway and may pinpoint therapeutic targets for Hh-based pathologies.