Drosophila tracheal cytonemes in long range FGF signaling

描述（由申请人提供）：这是一份申请K99/R00奖的申请材料，申请对象是加州大学旧金山分校的博士后Sougata Roy博士。Roy博士在生物化学，分子生物学，发育遗传学和细胞生物学方面经验丰富，并在他提出的目标中寻求先进成像技术的额外培训。Roy博士与他的主要导师Thomas B. Kornberg博士组建了一个指导团队，Thomas B. Kornberg博士是心血管研究所的教授，一位公认的发育生物学家，因其在遗传和hedgehog信号通路以及细胞素发现方面的基础性贡献而闻名；和两位顾问：David Agard博士，UCSF生物化学和生物物理系教授，世界知名，他在自动化EM断层扫描，3D重建和结构化照明方法开发方面的部分开创性工作；Ron Vale博士，UCSF细胞和分子药理学教授，因其对细胞骨架动力学和细胞内空间组织和运动的理解而闻名于世。罗伊博士的长期目标是了解细胞用于远距离交流的细胞和分子机制。他的建议侧重于细胞素的作用，这是一种信号丝状足，以及FGF（后生动物发育中的关键信号蛋白之一）在组织中移动并到达果蝇肺或气囊原基的机制。长期以来，人们一直认为信号蛋白如FGF在细胞外空间扩散并产生浓度梯度，这些梯度调节应答细胞的生长和模式。尽管多年来丝状足主要与迁移有关，但果蝇细胞素的发现和特征表明了信号蛋白运动的另一种模式-信号蛋白可能以受控的方式在直接接触点上传递。控制信号水平和靶点对正常和疾病状态都至关重要；一个说明性的例子是肺增生，在小鼠中可由FGF10信号的降低或升高引起。本研究旨在研究细胞素接触的性质，细胞素接触是信号蛋白从源细胞转移到靶细胞的位点，以及调节FGF运输和活性的机制。这些研究将采用果蝇遗传学、重组、RNAi筛选和最先进的显微镜技术，包括荧光显微镜、共聚焦显微镜、电子显微镜/三维电子扫描断层扫描、UCSF可用的结构照明显微镜（SIM）。拟议的研究将显著提高对细胞素介导的信号在肺发育过程中的机制和作用的理解，并有望在奖励结束前形成R01资助申请的基础。

DESCRIPTION (provided by applicant): 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.