Subhajit Poddar
Published: 2017
Total Pages: 124
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Viral infection of host cells induces the Type I interferon (IFN) response, which ischaracterized by the production of hundreds of IFN-stimulated genes (ISGs). Altogether, theseISGs function to induce an antiviral state, hindering or blocking various steps of the virallifecycle. Many individual ISGs have potent and broad antiviral functions. However eliminationof a single ISG does not completely abrogate protection, suggesting that other ISGs, althoughmoderate or moderate when considered alone, must work cooperatively to provide optimalantiviral activity.In order to identify and characterize novel ISGs, an attenuated strain of the alphaviruschikungunya (CHIKV-181/25) was tested against an shRNA library of 243 curated murine genesupregulated during IFN treatment. An attenuated CHIKV strain was used with the assumptionthat ISGs with moderate or low activity may be more easily identified due to the reducedpathogenicity 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 novelmurine ISGs that putatively restrict infection were identified from the CHIKV-181/25 screen andthe LACV screen, respectively.Although independent confirmation of many candidate antiviral ISG targets using bulkCRISPR lines is still ongoing, we were able to validate and characterize the antiviral role of oneof these targets, IFITM3, against alphaviruses in vitro and in vivo. Alphaviruses, which werepreviously thought to be unaffected by this ISG, exhibit reduced replication due to restriction byIfitm3 at the endosomal fusion stage of infection. Ifitm3-/- mice infected with CHIKV exhibitedgreater 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 cytokinesand chemokines in the ipsilateral foot. Splenic macrophages from Ifitm3-/- mice exhibited greaterlevels of viral antigen at 1 day after infection with CHIKV, and cultured bone marrow derivedmacrophages lacking Ifitm3 supported enhanced CHIKV replication. To test whether Ifitm3restricts encephalitic alphaviruses we infected WT and Ifitm3-/- mice with VEEV-TC83-A3G,and observed increased mortality and viral burden in Ifitm3-/-animals.