Functional Analysis of cilia in Hedgehog signaling and Gli activity regulation

从扁虫、果蝇到人类，动物的发育依赖于由Hedgehog(HH)家族信号蛋白介导的细胞之间的通讯。初级纤毛是细胞对HH作出反应所必需的，但目前尚不清楚它们是如何实现这一重要功能的。这个项目的重点是Gli2，它可以启动基因表达，以响应HH。首先，PI将采用分子方法来检验假设，即Gli2进入纤毛的运输与其进入细胞核的运输具有相同的机制。随后，PI将执行结构/功能分析，以确定Gli2蛋白中对其纤毛定位至关重要的氨基酸。最后，PI将创造一种基因工程小鼠，产生一种无法进入纤毛的Gli2蛋白。利用这只小鼠，PI将测试Gli2活动专门在纤毛中打开的假设。最近的报道将纤毛与许多其他信号蛋白(如WNT、BMP、PDGF等)联系在一起；因此，本研究不仅将促进对HH如何通过基于纤毛的信号来调节细胞行为的理解，而且还将为纤毛如何调节信息流以确保动物的正常发育提供更全面的见解。通过将研究材料应用于本科教学，鼓励本科生参与实验研究，为研究生提供参与教学活动的机会，实现研究与教育的融合。女性和少数族裔学生将通过NASA赞助的项目被招募参与这一项目。最后，PI将积极参与外联活动，如参加宾夕法尼亚州初级科学院活动(K-12教育)和为经济困难的第一代学生提供向上发展计划。

The development of animals ranging from flat worms and fruit flies to humans relies on communications between cells mediated by Hedgehog (Hh) family of signal proteins. The primary cilia, tiny 'hairs' on the surface of most of our cells, are needed for cells to respond to Hh, but it remains unclear how they achieve this important function. This project focuses on Gli2, which turns on gene expression in response to Hh. First, the PI will take a molecular approach to test the hypothesis that the transportation of Gli2 into the cilia share the same mechanism used for its transportation into the nucleus. Subsequently, the PI will perform a structure/function analysis to define amino acids in Gli2 protein critical for its ciliary localization. Finally, the PI will create a genetically engineered mouse that produces a Gli2 protein incapable of entering the cilia. Using this mouse, the PI will test the hypothesis that Gli2 activity is turned on specifically in the cilia. Recent reports linked the cilia to many other signal proteins (e.g. Wnt, Bmp, PDGF etc); hence this research will not only advance understanding of how Hh regulates cell behavior through cilia-based signaling, but also provide more general insight into how cilia regulate information flow to ensure normal development of an animal. The integration of research and education will be achieved by applying the research material to undergraduate teaching, encouraging undergraduate students to participate in lab research and providing opportunities for graduate students to be involved in teaching activities. Female and minority students will be recruited to participate in this project through NASA-sponsored programs. Finally, the PI will be actively involved in outreach activities such as participating in Pennsylvania Junior Academy of Science activities (K-12 education) and Upward-bound programs for first-generation, economically-challenged students.