Lkb1调控纤毛Sonic Hedgehog信号通路的分子机制研究

Lkb1是一个定位于初级纤毛的激酶，有报道其参与调控Sonic Hedgehog（SHH）信号通路，但Lkb1在SHH信号通路中功能及分子机制仍不明确。申请人的预实验结果表明：在3T3细胞中敲低Lkb1引起Gli3FL/R的比值升高；加入SHH信号通路的激动剂SAG，Gli2在纤毛顶部的集聚增加，Gli1的表达水平升高。外源表达Lkb1引起Gli3FL/R的比值降低；加入SAG后，Gli1的表达水平降低。由此，申请者人推测，Lkb1可以负调控SHH信号通路，可能的分子机制是Lkb1通过磷酸化修饰纤毛内的某个分子，参与Gli3的剪切过程，实现对SHH信号通路的精细调控，进而参与胚胎发育过程。为证实该假说，本项目拟通过分子生物学实验，细胞生物学实验以及小鼠模型，探讨Lkb1在SHH信号通路中的功能及其分子机制。该研究有望加深对SHH信号通路调控机理的理解，揭示Lkb1在胚胎发育过程中的功能。

Lkb1 is a well-known tumor suppressor gene and critical kinase involved in many physio-pathological processes, particularly embryonic development. Lkb1 can localize to the primary cilia, the specialized organelle that acts as the base for the Sonic Hedgehog (SHH) signaling pathway. Lkb1 is involved in the regulation of SHH signaling but the detailed molecular mechanisms and downstream cellular functions involving Lkb1 in SHH signaling remain unclear. Preliminary experimental results from the applicant show that the Gli3FL/R ratio which determines SHH pathway activity are increased when Lkb1 is knocked down. Moreover, Gli2, the SHH activator accumulates at the ciliary tip and the expression levels of the SHH responsive gene Gli1 are increased. Conversely, overexpression of Lkb1 decreases Gli3FL/R ratio and Gli1 levels. Notably, a kinase dead mutant of Lkb1 locates to the primary cilia but does not decrease Gli1 levels. Together this provides compelling evidence for the hypothesis that Lkb1 negatively regulates the SHH signaling pathway. The project will investigate the molecular mechanisms involved, identifying the downstream targets phosphorylated by Lkb1 in the cilia, and the associated processes that modulate the changes in the key SHH regulatory molecules. This will be achieved though the application of molecular and cellular biology experiments including mouse development models. The outcomes are expected to both enrich the understanding of the regulation of the SHH signaling pathway and reveal the function of Lkb1 during embryonic development.