Drosophila tracheal cytonemes in long range FGF signaling

项目摘要/摘要 这是该大学博士后苏加塔·罗伊博士的K99/R00奖项申请书 加利福尼亚州，旧金山。罗伊博士在生物化学、分子生物学、发育 遗传学和细胞生物学，并在他提出的目标中寻求先进成像技术的额外培训。 罗伊博士和他的主要导师托马斯·B·科恩伯格博士组建了一个指导团队 在心血管研究所，一位公认的发育生物学家，以其 在增强和Hedghog信号通路中的基本贡献以及对 和两位顾问：生物化学和生物物理系教授大卫·阿加德博士， 加州大学旧金山分校，世界知名，他在自动化EM方法开发方面的部分开创性工作 断层扫描、3D重建和结构照明；加州大学旧金山分校教授Ron Vale博士 细胞和分子药理学系，以其对认识的贡献而闻名世界 细胞骨架动力学和细胞内的空间组织和运动。 罗伊博士的长期目标是了解细胞用来 远距离交流。他的建议集中在细胞素的作用上，这是一种 以及后生动物的关键信号蛋白之一--成纤维细胞生长因子的作用机制 发育，穿过组织到达果蝇肺或气囊原基。一直以来都是 接受像成纤维细胞生长因子这样的信号蛋白在细胞外空间扩散以产生浓度梯度和 这些梯度调节反应细胞的生长和模式。尽管多年来丝状伪足一直是 主要与迁徙有关，果蝇细胞线的发现和特征表明 信号蛋白运动的另一种模型--信号蛋白可以在 在直接接触点控制时尚。控制信号水平和目标对于两者的正常 和疾病状态；一个说明性的例子是肺增生，在老鼠身上，它可能是由以下两种原因之一引起的 FGF10信号的降低或升高。本申请中提出的工作调查了 作为信号蛋白从源细胞转移到靶细胞的部位的细胞素接触，以及 调节成纤维细胞生长因子转运和活性的机制。这些研究将利用果蝇的遗传学， 重组工程、RNAi筛选和最先进的显微技术，包括荧光显微镜， 共焦显微镜、电子显微镜/三维电磁层析成像、结构照明显微镜 (SIM)在加州大学旧金山分校提供。 这项拟议的研究将显著提高对血管紧张素转换酶的机制和作用的理解。 细胞线粒体介导肺发育过程中的信号传递，预计将成为R01拨款的基础 在颁奖结束前提出申请。

PROJECT SUMMARY/ABSTRACT This is an application for a K99/R00 award for Dr. Sougata Roy, a post-doctoral fellow at the University of California, San Francisco. Dr. Roy is experienced in biochemistry, molecular biology, developmental genetics and cell biology and seeks additional training in advanced imaging techniques in his proposed goals. Dr. Roy has assembled a mentoring team with his primary mentor, Dr. Thomas B. Kornberg, Professor in the Cardiovascular Research Institute, a well-recognized developmental biologist and known for his fundamental contributions in the engrailed and Hedghog signaling pathways and for the discovery of cytonemes; and two advisors: Dr. David Agard, Professor in the Department of Biochemistry and Biophysics, UCSF, world-renowned, in part of his pioneering work with the development of methods for automated EM tomography, 3D reconstruction and structured illumination; and Dr. Ron Vale, Professor in the UCSF Department of Cellular and Molecular Pharmacology, world renowned for his contributions to understanding cytoskeletal dynamics and spatial organization and movement within cells. Dr. Roy's long-term goal is to understand the cellular and molecular mechanisms which cells use to communicate over long distances. His proposal focuses on the roles of cytonemes, which are a type of signaling filopodia, and on the mechanisms by which FGF, one of the key signaling proteins in metazoan development, moves across tissues and reaches Drosophila lung or air-sac primordium. It has long been accepted that signaling proteins like FGF diffuse in extracellular space to generate concentration gradients and that these gradients regulate growth and patterning of responding cells. Although for many years filopodia have been implicated principally in migration, the discovery and characterization of Drosophila cytonemes suggested an alternative model for movement of signaling proteins - that signaling proteins may be delivered in a controlled fashion at points of direct contact. Controlling signaling levels and targets is critical to both normal and disease states; an illustrative example is pulmonary hyperplasia, which in the mouse can result from either reduction or elevation of FGF10 signaling. The work proposed in this application investigates the nature of the cytoneme contacts that are the sites of transfer of signaling proteins from source to target cells, as well as the mechanisms that regulate FGF transport and activity. The studies will employ Drosophila genetics, recombineering, RNAi screen and state-of-the-art microscopy techniques, including fluorescent microscopy, confocal microscopy, Electron Microscopy/3-Dimension EM-tomography, structured illumination microscopy (SIM) available in UCSF. The proposed research will significantly improve understanding of the mechanism and role of cytoneme-mediated signaling during lung development and is expected to form the basis for an R01 grant application before the end of the award.