RNA interference (RNAi) is a conserved mechanism by which double-stranded RNA can specifically inactivate genes of matching sequence. RNAi has rapidly developed from a Nobel-prize winning discovery in the simple worm C. elegans to a therapeutic approach to silence human disease-causing genes that lack conventional medicines. However, fundamental aspects of RNAi remain unclear and they need to be understood to ensure safe and efficacious RNAi therapy. The continued relevance and significance of studies in the model organism C. elegans is underscored by the fact that the human counterpart of an RNA channel that imports RNA into C. elegans cells during RNAi is required for the import of RNAi-based drugs into human cells. The candidate presents a 5-year career development plan that aims to use C. elegans to gain fundamental insights into the transport of RNA between cells during RNA interference, while establishing an independent academic career at a research university. The candidate will build on his strong foundation in genetics and biochemistry to develop into an independent researcher in RNA transport under the mentorship of Dr. Craig Hunter, a pioneer and leader in the study of RNA transport between animal cells. The plan will be carried out in the Department of Molecular Biology at Harvard University, a leading institution in modern biology. Research in the mentor's lab led to the discovery of the conserved RNA channel SID-1 that is required for the import of RNAi-mediated silencing signals and the transport of signals between cells within a tissue. In a recent publication, the candidate reported the discovery that export of RNA from C. elegans tissues occurs through a regulated SID-1 independent mechanism. During the mentored phase, the candidate will: 1) Dissect the SID-1 dependent transport of RNA between cells within a tissue using advanced microscopy and examine how the SID-1 independent export of RNA from tissues is mediated by sid-3, a gene required for such export;and 2) examine the role of RNAi pathway proteins within a cell in generating RNAs transported from that cell. In addition to the mentor's laboratory, advanced microscopy for Aim1 will be carried out in the lab of the candidate's collaborator Dr. Xiaowei Zhuang, who is a pioneer in super-resolution microscopy. During the independent phase of the award, the candidate will analyze the roles of sexd-1 and sexd-2, two other genes discovered by the candidate that are required for inter-tissue export and will define a basic molecular pathway for export using the approaches and techniques acquired during the mentored phase Training in the complementary cell biological, genetic, and biochemical approaches while executing the above research plan will equip the candidate to establish a multi-faceted and rich research program as an independent investigator. Further, the proposed studies will reveal fundamental aspects of RNA transport during RNAi in C. elegans, which will impact the design of therapeutic RNAi approaches to human diseases.
RNAi-based drugs can specifically target disease-causing genes for which no conventional medicines are currently available. But, delivery of the drugs specifically and efficiently into diseased cells and tissues is a major barrier to RNA therapy. Understanding the mechanisms that control export of RNAi-mediated silencing signals from cells and the effect of such export on RNAi within these cells in C. elegans will provide valuable insights to overcome these barriers to therapeutic RNAi in humans.