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.
Proposed work will investigate a novel mechanism of regulated cell-cell communication. Preliminary work has shown that specialized filopodia (cytonemes) make contact between cells, even over long distances, and that these organelles mediate the regulated exchange of signaling proteins between producing and target cells. Understanding the mechanisms that these cytonemes use to move signaling proteins from one cell to another will give a new understanding of long distance signal transduction and will have profound implications for signaling in both normal development and disease.
