Thesis supervisors: Professor Andres Merits, Senior Research Fellow Eva Žusinaite.
Opponent: Diane E. Griffin, MD, PhD, University Distinguished Service Professor
W. Harry Feinstone Deprtment of Molecular Microbiology and Immunology, Johns Hopkins Bloomberg School of Public Health
Semliki Forest virus (SFV) is positive-sense RNA virus belonging to the genus Alphavirus in the family Togaviridae. This genus includes viruses pathogenic to a wide variety of animals, including humans, causing a spectrum of diseases that ranges from unpleasant flu-like illness and arthritis to fatal encephalitis. Laboratory strains of SFV have been utilized extensively in genetic engineering and they provide a well-characterized model system to investigate the pathogenesis of viral encephalitis. The most thoroughly studied strains of SFV are A7(74), SFV4, and L10. A7(74) is considered to be avirulent because the infection in adult mice is asymptomatic. In contrast, after intracerebral or high-dose intraperitoneal (i.p.) inoculation, L10 and SFV4 are both virulent and cause lethal encephalitis. However, following low-dose i.p. inoculation, SFV4 is incapable of reaching the brain, and the infected animals survive. Current thesis showed that phenotypic differences between SFV4 and L10 are determined by the charge of amino acid residues in viral glycoprotein E2. Positively charged amino acid residues in E2 facilitate the binding of SFV4 virions to heparan sulfate on the cell surface. This results in rapid clearance of virus from the blood and lower viremia, which in turn prevents the entry of the virus into the brain. We showed that phenotypic differences between A7(74) and L10 are caused by differences in the rate of nonstructural polyprotein processing and in the sequence of nonstructural protein 3 (nsP3). Slower processing of the polyprotein P123 1/2 cleavage site or the presence of L10 nsP3 enhance the ability of the virus replicase to induce interferon production, which correlates with virus ability to spread in the brain, suggesting a link between SFV neurovirulence and immunopathology. To further analyze the intricate interplay between the virus and the host, we developed a novel approach to determine the host proteins that colocalize with mature replication complexes of SFV. These data together help us better understand the neurovirulence of SFV. Furthermore, the methods and mechanisms discovered here can be applied in the research of medically important alphaviruses.