Regulation of Hedgehog Signaling by new cilium proteins

项目摘要 Hedgehog（Hh）信号广泛参与发育和癌症，包括髓母细胞瘤 (MB)小儿脑恶性肿瘤目前的治疗方案带来终身毁灭性的副作用， 幸存者，给患者家庭带来巨大负担。目前的Hh通路抑制剂受到挑战 耐药性和肿瘤复发。我们的目的是了解Hh的基本转导机制， 为Hh相关癌症的治疗提供新的线索。这一建议是基于我以前的 发现PDE 4D在特定亚细胞位点控制局部PKA活性以调节Hh信号传导。在 在过去的两年里，我们研究了Hh信号的控制，通过当地的PDE 4D-PKA在 中心体我们发现：1）PDE 4D 3通过myomegalin锚定在中心体上，局部抑制PKA 活性，2）用shRNA或CRISPR/Cas9敲除肌巨蛋白特异性地增加了PKA在肌巨蛋白水平的活性。 中心体，和3）这选择性地抑制Hh信号传导而不干扰其它PKA相关的细胞事件。 基于令人兴奋的结果和我们已经建立的创新工具，在目前的提案中，我们将扩大我们的 系统研究纤毛中局部PKA对Hh信号的调节。我们将1）定义 纤毛中从Smo到PKA的信号传导机制，Hh通路中的神秘步骤; 2）确定 选择性促进纤毛PKA对Hh信号传导和Hh相关肿瘤生长的影响;以及3）鉴定 通过基于邻近标记的蛋白质组学研究与Smo相关的信号转导子， Hh转导过程中的下游信号级联。我们的方法是创新的，因为它结合了切割 边缘技术来阐明Hh途径中长期存在的问题。它的重要性被强调为 期望它将突出治疗小儿脑肿瘤的新方法，副作用更少。 同样重要的是，这笔研究资金将使我能够让第一代大学生参与 加州大学默塞德分校的生物医学研究，以多样化和加强在经济贫困的学术研究 加州中央谷。

PROJECT SUMMARY Hedgehog (Hh) signaling is widely involved in development and cancers, including medulloblastoma (MB), the malignant pediatric brain tumor. Current treatment regimens bring life-long devastating side effects to survivors, generating a huge burden to the patients’ family. The current Hh pathway inhibitors are challenged by drug resistance and tumor relapse. We aim to understand the basic transduction mechanisms of the Hh pathway in order to provide new clues to treat Hh-related cancers. This proposal is based on my previous discovery that PDE4D controls local PKA activity at specific subcellular sites to regulate Hh signaling. In the past two years of this grant, we have studied the control of Hh signaling by local PDE4D-PKA at the centrosome. We found that 1) PDE4D3 is anchored to the centrosome by myomegalin to locally inhibit PKA activity, 2) knocking out myomegalin with shRNA or CRISPR/Cas9 specifically increases PKA activity at the centrosome, and 3) this selectively inhibits Hh signaling without interfering other PKA-related cellular events. Based on the exciting results and the innovative tools we have built, in the current proposal we will expand our research to systemically investigate the regulation of Hh signaling by local PKA in the cilium. We will 1) define the signaling mechanism from Smo to PKA in the cilium, a mysterious step in the Hh pathway; 2) determine the effects of selectively promoting ciliary PKA on Hh signaling and Hh-related tumor growth; and 3) identify the signaling transducers associated with Smo via proximity labeling-based proteomics to thoroughly illuminate its downstream signaling cascade during Hh transduction. Our approach is innovative because it combines cutting edge techniques to elucidate the long-standing questions in the Hh pathway. Its significance is underlined by the expectation that it will highlight new approaches to treat pediatric brain tumor with fewer side effects. Equally importantly, this research funding will allow me to engage first generation college students in biomedical research at UC Merced to diversify and strengthen academic research in the economically deprived California Central valley.