Egelman, Edward H $372,504/yr. 09/15/14 – 08/31/18
Public Health Relevance:
This project is aimed at understanding the structures of a range of nucleoprotein complexes. These complexes include viral capsids, filaments that form as part of the innate immune system to recognize viral RNA, and assemblies involved in DNA repair. Almost all of these complexes are directly related to human health, and could be important in designing new strategies to deal with bacterial and viral pathogenesis, as well as to better understand the breakdowns of DNA repair that lead to cancer.
This project, currently in its 27th year, has been aimed at understanding the structure of many nucleoprotein complexes, particularly those involved in homologous genetic recombination. Almost all of the proposed projects will depend upon our newly installed Titan Krios cryo-EM. This microscope, combined with our extensive experience in the area of heterogeneous helical polymers, puts us in a unique position to accomplish the stated aims:
1) MDA5-RNA Filaments – The innate immune system provides a defense against many microbial structures, but is poorly understood. Extending our preliminary results will have a significant impact, and high-resolution EM provides the best way to study both the protein and RNA in these filaments.
2) RIG-IRNA Filaments – We can directly address the controversy surrounding whether this protein forms filaments, and understand the similarities and differences with the MDA5 system.
3) Binding of HIV NC to Actin –Preliminary results show a highly specific binding, and the structural studies that we propose may lead to entirely new targets for preventing HIV transmission.
4) Cas7 Filaments – Like MDA5 and RIG-I, Cas7 is part of an innate immune system, but in archaea.
5) Xrcc4-Cernunnos Filaments – We have worked for many years on proteins and filaments involved in homologous recombination. These filaments are part of the Non-Homologous End Joining pathway for DNA repair, and are of great interest and significance to understanding failures in DNA repair which lead to genomic instability and cancer.
6) Viral Capsid Tubes –We will continue our work on helical tubes formed from both the P22 and HIV capsid proteins. With P22 these tubes will be modified to contain RNA. Both projects, when extended to high resolution, should yield new insights into the ability of capsid hexamers to switch conformations.