Date of Award
Doctor of Philosophy (PhD)
The journey from plasma membrane to nuclear pore is a critical step in the lifecycle of DNA viruses, many of which must successfully deposit their genomes into the nucleus for replication. Viral capsids strategically navigate this vast distance (and all subsequent lifecycle steps) through the coordinated hijacking of a number of cellular proteins subsequently termed host factors. Given the virus’ dependence on these proteins, host factors therefore represent valuable targets for therapeutic interventions. Still, the identity and function of many of these factors remains unknown. In this body of work, I will detail our own journey from initial identification to comprehensive characterization of one such host factor, Mindbomb 1 (MIB1), in the context of adenovirus infection. Adenoviruses (AdVs) are widespread and highly contagious DNA viruses that can cause severe respiratory illness in children and immune-compromised individuals. An initial genome-wide loss-of-function screen to identify host factors for this virus revealed MIB1, an E3 ubiquitin ligase best known for its role in neurodevelopment, as critical for AdV infectivity. In a series of mechanistic studies centered on the earliest stage of infection, we observed that in the absence of MIB1, viral capsids successfully traffic to the proximity of the nucleus but ultimately fail to deliver their genomes within. AdV infection is dependent on MIB1’s primary action as an E3 ubiquitin ligase — to carry out ubiquitination, a posttranslational modification widely capitalized upon by viruses for rapid manipulation of the host environment via altered protein localization, activity or turnover. Our work suggests that in the immediate vicinity of the nucleus, MIB1 may be required for the proteasomal degradation of one or more negative regulators of AdV infection. To identify this relevant MIB1-ubiquitination target, we turned to complementary proteomic approaches to determine proteins proximal to MIB1 upon AdV infection and those differentially ubiquitinated in its presence or absence. Using these unbiased approaches, we corroborated previous reports of MIB1 as a core component of centriollar satellites, dynamic structures localized to the intervening distance between centrosome and nucleus. Furthermore, both proteomic approaches independently pointed to an understudied yet evolutionarily-conserved role for MIB1 in regulating RNP granules and cytoskeleton within the perinuclear environment. Understanding the full relevance of these MIB1-regulated pathways to AdV infection and identifying the specific ubiquitination target responsible will be natural extensions of this work to fully dissect the mechanism of MIB1-mediated viral genome delivery. Together, this work highlights yet another creative way in which viruses recruit host cell machinery to facilitate their replication with the potential to inform the design of new antiviral treatments and emerging adenoviral vector-based therapies.
Sarbanes, Stephanie, "A Role for Mindbomb 1 in Adenovirus Genome Delivery" (2020). Student Theses and Dissertations. 704.