Coronavirus Nonstructural Protein Interactions with Endoplasmic Reticulum Proteostasis Factors Mediate Infection

Abstract

My thesis work examines how coronaviruses hijack host cell proteins and stress response pathways during infection. Coronaviruses encode sixteen nonstructural proteins (nsps), which interact with host proteins to modulate host membranes and stress response pathways to mediate viral replication. An important question with the emergence of the COVID-19 pandemic was how do SARS-CoV-2 nsps interact with host proteins to mediate infection? And are these interactions similar or different to the strategies employed by previously known CoVs, like SARS-CoV, MERS-CoV, hCoV-OC43, hCoV-229E, and Murine Hepatitis Virus (MHV)? Towards this end, I used quantitative proteomics to comparatively profile the interaction network of several CoV nsp homologs (nsp2, nsp3, nsp4), discovering a diverse array of both conserved and unique host interactors. I then applied a functional genetic screen to identify interactors with important roles in CoV infection. I found several conserved, pro-viral interactors associated with protein biogenesis, including the glycoprotein quality control factor Malectin (Mlec). Further testing reveals that Mlec promotes early nsp production and mediates replicase formation through the glycoprotein biogenesis pathway. In addition, I have also characterized the role of nsps in modulating an important stress response pathway, the Unfolded Protein Response (UPR), which senses and mitigates build-up of misfolded proteins in the endoplasmic reticulum to maintain protein homeostasis. Using quantitative proteomics, I found that SARS-CoV-2 nsp3 and nsp4 work in concert to tune the UPR in a temporal manner that may increase viral protein folding capacity while limiting potentially pathogenic effects caused by chronic UPR activation. Together, my thesis work reveals new strategies by which CoV nsps can co-opt host proteins and regulatory systems to mediate viral infection. These dependencies could potentially be exploited for the design of new pan-CoV antiviral therapies in preparation for future CoV outbreaks.