. The goal of this administrative supplement request is to provide a state-of-the- art microscope for cell biological studies of lysosomal sorting for R01GM120735. The requested instrument will be shared equally with Drs. Blake Hill (R01GM067180.) and Matt Scaglione (R35GM119544) for their cell biological studies in mitochondria fission and proteostasis, respectively. The herpesviruses have co-evolved with their hosts for hundreds of millions of years. As our adaptive immune systems evolved, so did the mechanisms of immune evasion encoded by herpesviruses. Because herpesviruses remain latent or persistent within their host throughout life, the herpesviruses have evolved a unique set of strategies to help them contend with the lifelong host immune response. This application?s central premise is that the herpesviruses, in their evolution, have performed a 100 million-year-long genetic screen to undermine host defense mechanisms, and in so doing, can illuminate cell biological processes that we may not yet understand; we seek to take advantage of their evolution to discover new concepts in cell biology. The long term goal of this project is to understand the underlying cell biological mechanisms by which human herpesviruses-6 and -7 (HHV-6/7) escape host defense mechanisms. The focus of this proposal is to understand how HHV-7 U21 reroutes class I MHC molecules to lysosomes, and in so doing, understand the cellular pathway usurped by U21. Our preliminary data indicate that U21 employs an unknown sorting mechanism that seems to defy convention - almost certainly utilizing a novel cellular means of lysosomal sorting.
In Aim 1, we assess the structure and association of U21 with itself, as a homotetramers, and determine whether homooligomerization of U21 is necessary its ability to bind to and reroute class I MHC molecules to lysosomes, and use RUSH to examine the synchronous trafficking of U21 and class I molecules to lysosomes. We ultimately seek a temporal and mechanistic view of how U21 undermines the cellular lysosomal trafficking pathway, and how this lysosomal trafficking pathway functions in the absence of U21. A better understanding of this process should contribute to our understanding of basic underlying principles of lysosomal trafficking, and also lead to a more thorough comprehension of how these viruses have such great success in evading the host immune response.
Human herpesviruses-6 and -7 infect essentially 100% of the world's population before the age of 5 and then remain latent or persistent in their host throughout life. As such, these viruses are among the most pervasive and stealthy of all viruses. They must necessarily excel at escaping immune detection throughout the life of the host. HHV-6 and -7 both encode a protein, U21, that binds to and reroutes the host immune defense molecules class I MHC and NK activating ligands to lysosomes, where they are degraded. This project seeks to identify the cellular protein(s) usurped by U21 as it reroutes class I MHC molecules to the lysosomal compartment. It also seeks to assess the molecular and structural features of U21 that allow it to function.
