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David J. Miller

Assistant Professor
M.D., Ph.D, Mayo Clinic, 1996

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Host-Pathogen Interactions in Viral RNA Replication

Our laboratory investigates the molecular and cellular mechanisms responsible for viral RNA replication complex assembly, function, and regulation, and the interactions between viral RNA replication and the innate cellular antiviral responses. The clinical manifestations and outcomes of viral infections are dependent upon dynamic and complex host-pathogen interactions. During the viral life cycle these interactions occur at several steps, including virus attachment, transcription and viral protein synthesis, genome replication and encapsidation, and eventually the release of infectious virus particles. The importance of particular interactions is highlighted by the demonstration that viruses frequently use common pathways to accomplish essential steps in their replication cycle. For example, positive-strand RNA viruses universally assemble their viral RNA replication factories as macromolecular complexes that are physically associated with host intracellular membranes. In addition, positive-strand RNA viruses often produce double-stranded RNA during their replication cycle, and thus can trigger one of several innate intracellular antiviral responses.

We study the host-pathogen interactions involved in viral RNA replication using two positive-strand RNA viruses, Flock House virus (FHV) and western equine encephalitis virus (WEEV). FHV is a model insect pathogen that has one of the smallest known genomes of any animal RNA virus, and normally assembles its RNA replication complexes on outer mitochondrial membranes. One particularly useful aspect of FHV is its robust replication in several genetically-tractable hosts, including the yeast Saccharomyces cerevisiae and cells derived from the fruit fly Drosophila melanogaster. These features make FHV an extremely useful model system to study the molecular interactions between viral RNA replication components and cellular factors. Our research focuses on the mechanisms of intracellular targeting, trafficking, and assembly of viral replication complexes, and the structural and functional consequences of the interactions between viral RNA replication complexes and host intracellular membranes.

We also study the intracellular pathogenesis of WEEV, which is an arbovirus responsible for central nervous system infections in humans, and is also listed as a CDC and NIAID Category B potential bioterrorism agent. Our research focuses on WEEV RNA replication complex assembly, structure, and function, and its impact on the innate intracellular antiviral response in both the insect vector and the vertebrate target cell. As obligate intracellular pathogens, viruses must survive within an intrinsically hostile environment, and thus often use counter defense strategies to suppress or evade the innate immune response. Arboviruses must also survive and flourish in both vector and target host cells, which often employ significantly different innate antiviral responses. Understanding the mechanisms whereby arboviruses interact with the host to suppress or evade cellular defenses, and the impact of viral RNA replication complex assembly and function on these interactions, represents a crucial area of investigation in viral pathogenesis, and may identify potential novel targets for antiviral therapies.

Selected Publications:

Miller, D.J., Schwartz, M.D., and Ahlquist, P. Flock House virus RNA replicates on the outer mitochondrial membrane of Drosophila cells. J. Virol., 75:11664-11676, 2001.

Miller, D.J. and Ahlquist, P. Flock House virus RNA polymerase is a transmembrane protein with amino-terminus sequences sufficient for mitochondrial localization and membrane insertion. J. Virol., 76:9856-9867, 2002.

Miller, D.J., Schwartz, M.D., Dye, B.T., and Ahlquist, P. Engineered retargeting of viral RNA replication complexes to an alternative intracellular membrane. J. Virol. 77:12193-12202, 2003.

Dye, B.T., Schell, K., Miller, D.J., and Ahlquist, P. Detecting protein-protein interactions in live yeast by flow cytometry. Cytometry, 63A:77-86, 2005.

Kampmueller, K.M. and Miller, D.J. The cellular chaperone heat-shock protein 90 facilitates Flock House virus RNA replication in Drosophila cells. J. Virol. 79:6827-6837, 2005.

Dye, B.T., Miller, D.J., and Ahlquist, P. Self-interaction of nodavirus RNA replicase protein A in vivo. J. Virol., in press.