Biochemical mechanisms of Hedgehog signal transduction through primary cilia

项目摘要 信号分子的时空调节对于发育和成体组织的稳态是至关重要的。 事实上，信号传导事件的错误激活或错误定位可导致先天性疾病和癌症， 了解细胞中的信息传递是如何调节的，对于理解和改善人类健康至关重要。 健康刺猬途径在后生动物中是保守的，它控制细胞增殖， 分化、迁移和体内平衡。在脊椎动物中，Hedgehog信号通过初级 从大多数细胞表面伸出的纤毛，包括由Hedgehog错误激活驱动的癌症细胞 通路纤毛是脊椎动物Hedgehog信号所必需的，其原因尚不清楚。我们的中央 假设是睫状体微环境使蛋白质和脂质相互作用成为必要， Hedgehog信号转导我们的实验室已经确定了纤毛相关脂质的重要作用， 平滑，积累平滑纤毛，并激活刺猬途径。当刺猬 通路关闭，纤毛中的平滑积累和活性被Patched1抑制。如何修补1和 Smoothened在纤毛中的积累和功能，以及Patched1如何抑制Smoothened，目前还知之甚少 尽管越来越多的人认识到Hedgehog途径的这些活性在健康和疾病中至关重要。 为了解决我们对人类生物学理解的这些关键差距，本提案的目标是 定义蛋白质相互作用是必要的补丁1和平滑的积累和活动， 初级纤毛，并确定Patched1是否通过纤毛脂质的区室化抑制Smoothened。 最近，我们已经开始意识到，刺猬途径可以直接从大脑中收集信息。 纤毛微环境我们发现的睫状体脂质，指定刺猬途径输出建立了一个新的 范式的信息输入到刺猬转录程序，其中纤毛的变化， 微环境重新编程刺猬反应。在本建议中，我们将系统地研究 纤毛中蛋白质和脂质相互作用的动态库，其指定了正常和 肿瘤细胞我们将利用蛋白质组邻近标记质谱法的最新技术进展， 脂质组学质谱和功能基因组学来解释细胞状态的高维景观 Hedgehog pathway activation激活。 这项提案的长期目标是了解刺猬信号的机制 转导足够好，以开发Hedgehog相关癌症的新疗法。为此，数据 本提案产生的结果将建立一个基本原理，以在体内和样品中询问这些机制， Hedgehog相关癌症的人类患者。更广泛地说，这一建议将阐明为什么纤毛是 Hedgehog信号传导所需的信息以及细胞中信息传递的调节方式，是一个难以捉摸的基本问题。 人类生物学的问题。

Project summary Spatiotemporal regulation of signal molecules is critical for development and adult tissue homeostasis. Indeed, misactivation or mislocalization of signaling events can cause congenital disorders and cancers, and understanding how information transfer is regulated in cells is essential for understanding and improving human health. The Hedgehog pathway is conserved across metazoan animals where it controls cell proliferation, differentiation, migration, and homeostasis. In vertebrates, Hedgehog signals are transduced through primary cilia that project from the surface of most cells, including cells in cancers driven by misactivation of the Hedgehog pathway. Cilia are required for vertebrate Hedgehog signaling for reasons that remain unknown. Our central hypothesis is that the ciliary microenvironment enables protein and lipid interactions that are necessary for Hedgehog signal transduction. Our lab has identified an important role for cilia-associated lipids that bind to Smoothened, accumulate Smoothened to cilia, and activate the Hedgehog pathway. When the Hedgehog pathway is off, Smoothened accumulation and activity in cilia is inhibited by Patched1. How Patched1 and Smoothened accumulate and function in cilia, and how Patched1 inhibits Smoothened, are poorly understood despite increasing recognition that these activities of the Hedgehog pathway are critical in health and disease. To address these key gaps in our understanding of human biology, the objectives of this proposal are to define the protein interactions that are necessary for Patched1 and Smoothened accumulation and activity in primary cilia, and to determine if Patched1 inhibited Smoothened through compartmentalization of ciliary lipids. Recently, we have begun to appreciate that the Hedgehog pathway can gather information directly from the ciliary microenvironment. Our discovery of ciliary lipids that specify Hedgehog pathway output establishes a new paradigm for information input onto the Hedgehog transcriptional program, one in which changes in the ciliary microenvironment reprogram the Hedgehog response. In this proposal, we will systematically examine the dynamic repertoire of protein and lipid interactions in cilia that specify the Hedgehog pathway across normal and tumor cells. We will leverage recent technical advances in proteomic proximity-labeling mass spectrometry, lipidomic mass spectrometry, and functional genomics to interpret high-dimensional landscapes of cell states induced by Hedgehog pathway activation. The long-term goal of this proposal is to understanding mechanisms underlying Hedgehog signal transduction well enough to develop new treatments for Hedgehog-associated cancers. To do so, the data generated by this proposal will establish a rationale to interrogate these mechanisms in vivo and in samples from human patients with Hedgehog-associated cancers. More broadly, this proposal will elucidate why cilia are required for Hedgehog signaling and how information transfer is regulated in cells, an elusive and fundamental question of human biology.