Date of Award

Summer 8-15-2017

Author's School

Graduate School of Arts and Sciences

Author's Department

Biology & Biomedical Sciences (Immunology)

Degree Name

Doctor of Philosophy (PhD)

Degree Type

Dissertation

Abstract

Viral infection of host cells induces the Type I interferon (IFN) response, which is

characterized by the production of hundreds of IFN-stimulated genes (ISGs). Altogether, these

ISGs function to induce an antiviral state, hindering or blocking various steps of the viral

lifecycle. Many individual ISGs have potent and broad antiviral functions. However elimination

of a single ISG does not completely abrogate protection, suggesting that other ISGs, although

moderate or moderate when considered alone, must work cooperatively to provide optimal

antiviral activity.

In order to identify and characterize novel ISGs, an attenuated strain of the alphavirus

chikungunya (CHIKV-181/25) was tested against an shRNA library of 243 curated murine genes

upregulated during IFN treatment. An attenuated CHIKV strain was used with the assumption

that ISGs with moderate or low activity may be more easily identified due to the reduced

pathogenicity of the virus. In addition, the orthobunyavirus LaCrosse (LACV) was also tested, as there have been no large scale ISG screens using this pathogen. A total of 21 and 30 novel

murine ISGs that putatively restrict infection were identified from the CHIKV-181/25 screen and

the LACV screen, respectively.

Although independent confirmation of many candidate antiviral ISG targets using bulk

CRISPR lines is still ongoing, we were able to validate and characterize the antiviral role of one

of these targets, IFITM3, against alphaviruses in vitro and in vivo. Alphaviruses, which were

previously thought to be unaffected by this ISG, exhibit reduced replication due to restriction by

Ifitm3 at the endosomal fusion stage of infection. Ifitm3-/- mice infected with CHIKV exhibited

greater swelling of the ipsilateral foot at peak days of pathology. Higher viral titers in the spleen,

serum and ipsilateral foot were seen at 1 day after infection, coinciding with increased cytokines

and chemokines in the ipsilateral foot. Splenic macrophages from Ifitm3-/- mice exhibited greater

levels of viral antigen at 1 day after infection with CHIKV, and cultured bone marrow derived

macrophages lacking Ifitm3 supported enhanced CHIKV replication. To test whether Ifitm3

restricts encephalitic alphaviruses we infected WT and Ifitm3-/- mice with VEEV-TC83-A3G,

and observed increased mortality and viral burden in Ifitm3-/-

animals.

Language

English (en)

Chair and Committee

Michael S. Diamond

Committee Members

Deborah Lenschow, Daved Fremont, Marco Colonna, Adrianus C. Boon,

Comments

Permanent URL: https://doi.org/10.7936/K7MC8ZDM

Download

Share

COinS