Center for Structural Biology of HIV RNA

Core 3. Imaging Summary The purpose of the CRNA Imaging Core is to provide CRNA investigators with access to cutting edge imaging approaches and expertise in both light microscopy and cryo-electron microscopy (cryo-EM). The Core will collaborate with CRNA investigators to use high-resolution light microscopy to localize and track HIV-1 RNA- protein complexes in their cellular context and to use single particle cryo-electron microscopy (cryo-EM) to determine structures of RNAs, RNA-protein complexes, and HIV-related protein complexes. The Core will work to bridge the resolution gap between the two approaches by integrating live cell and superresolution light microscopy approaches, facilitated by the rapidly improving methods in cryo-electron tomography (cryo-ET). The overall goal is to be able to determine 3D structures of HIV RNA-protein complexes both in vitro and in their cellular context. The Core lab is co-led by two PIs. The component at the University of Michigan is focused on cryo-EM and is directed by PI Melanie Ohi. The lab at the University of Wisconsin, run by PI Nathan Sherer, will provide resources and expertise in light microscopy approaches. The University of Michigan Core lab will serve as a repository for materials and protocols. The Core team will focus on three aims that include utilizing single particle cryo-EM approaches to determine structures of RNAs, RNA-protein complexes and HIV-1-related protein complexes; developing effective “sample-to-structure” pipelines for efficient analysis of samples that will leverage the Core’s ability to image at resolutions ranging from ~50 Å to sub-3 Å; and moving towards using correlative light and electron microscopy (CLEM) and cryo-ET to detect and determine structures inside cells. In support of its training mission, the Core will also develop innovative educational programs for CRNA trainees seeking to become independent practitioners of these methods and techniques.