Characterization of Innate Immune Mechanisms Against RVFV and a Potential Target for Antivirals
Head, Jennifer 1980-
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Rift Valley fever virus (RVFV) is a mosquito-borne zoonotic pathogen, which affects both humans and livestock. RVFV belongs to the Bunyaviridae family, genus Phlebovirus and consists of a tripartite genome (L-, M- and S- segments). NSs, encoded in the S- segment promotes degradation of PKR and general transcription shutoff including IFN-β mRNA synthesis. Currently, no effective measures have been developed to prevent or treat RVFV disease and very little is understood about what takes place during the initial stages of RVFV infection. The objective of this series of studies was to characterize RVFV infection in human dendritic cells (DC) and to develop a dominant-negative method to attenuate RVFV NSs protein as a treatment for RVFV. Human DC displayed low permissiveness to RVFV but initiated high levels of pro-inflammatory mediator secretion in response to the virus. DCs infected with rMP12-C13type virus, encoding a non-functional NSs protein, were able to initiate increased levels of maturation marker presentation of CD40, CD86 and CD83 and were also able to induce higher levels of proinflammatory molecules while rMP-12 and rMP12-delNSm/78 viruses (expressing functional NSs protein but lacking 78 kDa protein and NSm) did not. We found that NSm does not play a major role in DC activation during RVFV infection. We then attempted to develop a dominant-negative strategy to inhibit RVFV NSs. NSs forms filamentous inclusions within the nucleus and self-associates at the C-terminus of the NSs protein. We characterized 11 different MP-12 mutants, each encoding a 17-25 amino acid truncation in NSs. Unexpectedly, each of the mutants lacked NSs functions to inhibit innate immune responses due to the truncations. Although the NSs proteins were still able to accumulate within the cells, they did not interact with MP-12 NSs when co-expressed in cells. In cells infected with MP-12 and treated with in vitro synthesized RNA encoding truncated nonfunctional NSs, MP-12 NSs was still able to inhibit IFN-β and was able to degrade PKR protein, indicating that co-expression of non-functional NSs does not exhibit dominant-negative phenotypes.