Date of Award

2013

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Immunology

First Advisor

Sara Cherry

Abstract

Arthropod-borne viruses (arboviruses) are an important class of emerging pathogens that cause mortality and morbidity worldwide. As obligate intracellular parasites with limited coding capacity, viruses must hijack host factors to replicate while evading host detection. To date, no specific therapeutic interventions exist for arboviruses and most lack FDA approved vaccines. This is in part due to a lack of understanding of viral-host interactions. To identify host factors that impact infection, we performed a genome-wide RNAi screen in Drosophila and identified 131 genes that affected infection of the mosquito-transmitted bunyavirus Rift Valley Fever virus (RVFV). Dcp2, the catalytic component of the mRNA decapping machinery, and two decapping activators, DDX6 and LSM7, were antiviral against disparate bunyaviruses in both insect cells and adult flies. Bunyaviruses 5' cap their mRNAs by `cap-snatching' the 5' ends of poorly defined host mRNAs. We found that RVFV cap-snatches the 5' ends of Dcp2 targeted mRNAs, including cell cycle related genes. Loss of Dcp2 allows increased viral transcription, while ectopic expression of Dcp2 impedes transcription. Furthermore, arresting cells in late S/early G2 led to increased Dcp2 mRNA targets and increased RVFV replication. Therefore, RVFV competes for the Dcp2-accessible mRNA pool, which is dynamically regulated and can present a bottleneck for viral replication.

I extended these studies to mammalian cells and I found that the two known human decapping enzymes, DCP2 and NUDT16, restrict RVFV replication. Since depletion of either gene impacted replication, this suggests that DCP2 and NUDT16 are non-redundant. In human cells, I found that RVFV predominately cap-snatches from mRNAs associated with translation, and the stability of these mRNAs is regulated by decapping; furthermore, instability of these mRNAs is triggered by RVFV infection. I hypothesize that translationally-associated genes, including ribosomal protein mRNAs, are selectively degraded during this response to limit translation to prevent viral replication. These data suggest that a particular functional class of mRNAs (translation-associated) can be coordinately regulated at the level of mRNA stability by decapping, and that this may be used as a mechanism by cells to selectively regulate gene expression.

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