Date of Award

Winter 1-13-2026

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Microbiology and Immunology

First Advisor

Thomas Gallagher

Abstract

Coronavirus (CoV) structural proteins congregate at the ER-Golgi intermediate compartments (ERGIC) prior to virus budding into the organelle lumen. Subsequent trafficking and fusion of virus-containing, ERGIC-derived vesicles then release nascent viruses from infected cells. Our findings in this dissertation illuminate mechanisms by which CoV structural proteins coalesce at the virus assembly and budding sites. The cytoplasmic tails of all CoV spike (S) glycoproteins contain variants of a consensus KxKxx coatomer-binding motif. Host coatomer complexes facilitate retrograde trafficking of client cargo proteins via recognition of such motifs. Utilizing reverse genetics methodologies, we introduced variations in the S coatomer motif in a prototype mouse hepatitis coronavirus (MHV) model. Viruses with altered coatomer affinities were debilitated and rapidly acquired suppressor mutations. These variations had profound effects on S intracellular localization, and subsequent interaction with other viral structural proteins, yet particles were released from infected cells. Secreted viral particles with altered motifs had reduced specific infectivities, which could be traced to sub-optimal S protein incorporation into virions. Our findings point to precisely evolved affinities between CoV S and host coatomer complexes, such that the requisite levels of S proteins are trafficked to ERGIC, where they integrate with other viral proteins for efficient virion assembly. Findings from this work provide a better understanding of CoV S intracellular trafficking and highlight their previously underappreciated influence on particle assembly.

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