Date of Award

2015-01-01

Degree Name

Doctor of Philosophy

Department

Biological Sciences

Advisor(s)

Kyle L. Johnson

Abstract

Positive-strand RNA viruses amplify their genomes in membrane-bound structures associated with intracellular membranes and organelles called replication complexes (RCs). Here, we begin to elucidate mechanisms of Nodamura virus (NoV; family Nodaviridae) RC assembly. The literature reports that NoV-infected muscle tissue exhibits mitochondrial aggregation and rearrangement of mitochondrial structure, leading to disorganization of the muscle fibrils. However, the molecular basis for this pathogenesis and the role of mitochondria in NoV infection remained unclear until now. We tested the hypoThesis that NoV establishes RCs in association with mitochondria in cultured mammalian cells at physiological temperature. We used immunofluorescence confocal microscopy and biochemical methods to determine that the aggregated mitochondria represent the sites of NoV RCs. We determined that the NoV RdRp (protein A; NA) uses two membrane-associated regions to localize to mitochondria as an integral membrane protein. Cells expressing NA exhibit mitochondrial morphologies similar to that described for diabetes, neurodegenerative disease and cardiovascular disease. Mitochondrial morphology is controlled by a cyclical mechanism referred to as mitochondrial dynamics, by which mitochondria cycle between two distinct forms: organelle and tubular. The tubular form resembles the clustered phenotype we observe in the presence of NA. This led us to hypothesize that NA may disrupt mitochondrial dynamics, leading to accumulation of clustered mitochondria for the purpose of establishing viral RNA replication complexes.

Language

en

Provenance

Received from ProQuest

File Size

222 pages

File Format

application/pdf

Rights Holder

Vincent Ulysses Gant

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