Abstract
Rift Valley fever virus (RVFV), belongs to genus Phlebovirus of the family Bunyaviridae, causes high rates of abortion and fetal malformation in infected ruminants as well as causing neurological disorders, blindness, or lethal hemorrhagic fever in humans. RVFV is classified as a category A priority pathogen and a select agent in the U.S., and currently there are no therapeutics available for RVF patients. NSs protein, a major virulence factor of RVFV, inhibits host transcription including interferon (IFN)-β mRNA synthesis and promotes degradation of dsRNA-dependent protein kinase (PKR). NSs self-associates at the C-terminus 17 aa., while NSs at aa.210–230 binds to Sin3A-associated protein (SAP30) to inhibit the activation of IFN-β promoter. Thus, we hypothesize that NSs function(s) can be abolished by truncation of specific domains, and co-expression of nonfunctional NSs with intact NSs will result in the attenuation of NSs function by dominant-negative effect. Unexpectedly, we found that RVFV NSs truncated at aa. 6–30, 31–55, 56–80, 81–105, 106–130, 131–155, 156–180, 181–205, 206–230, 231–248 or 249–265 lack functions of IFN–β mRNA synthesis inhibition and degradation of PKR. Truncated NSs were less stable in infected cells, while nuclear localization was inhibited in NSs lacking either of aa.81–105, 106–130, 131–155, 156–180, 181–205, 206–230 or 231–248. Furthermore, none of truncated NSs had exhibited significant dominant-negative functions for NSs-mediated IFN-β suppression or PKR degradation upon co-expression in cells infected with RVFV. We also found that any of truncated NSs except for intact NSs does not interact with RVFV NSs even in the presence of intact C-terminus self-association domain. Our results suggest that conformational integrity of NSs is important for the stability, cellular localization and biological functions of RVFV NSs, and the co-expression of truncated NSs does not exhibit dominant-negative phenotype.
Highlights
Rift Valley fever virus (RVFV) belongs to genus Phlebovirus of the family Bunyaviridae, and is a mosquito-borne zoonotic pathogen which causes Rift Valley fever (RVF)
Since NSs can self-associate and form filamentous inclusion bodies in infected cells [65,77], we hypothesized that co-expression of nonfunctional NSs with the C-terminal self-association domain in cells infected with RVFV allows incorporation of such nonfunctional NSs into the NSs filament, and attenuates a part of NSs functions
Our results were not as expected; 1) most of truncated NSs localized at cytoplasm, 2) all of truncated NSs did not accumulate as much as parental NSs in cells, and 3) none of truncated NSs significantly interfered with MP-12 NSs functions
Summary
Rift Valley fever virus (RVFV) belongs to genus Phlebovirus of the family Bunyaviridae, and is a mosquito-borne zoonotic pathogen which causes Rift Valley fever (RVF). The first recognized outbreak of RVF occurred in Kenya in 1930 [6], and RVFV has spread from endemic region in sub-Saharan Africa into Egypt [7], Madagascar and the Arabian Peninsula [8,9,10,11,12]. RVFV is a risk group 3 pathogen, Category A pathogen and an overlap select agent by the CDC/USDA [16]. The handling of wild-type (wt) RVFV within the U.S requires BSL3+ or BSL4 facilities. Live-attenuated MP12 vaccine strain is excluded from select agent rule, and handled at BSL2 level. MP-12 encodes for functional NSs protein, which is useful for the analyses of various NSs functions at BSL2 level [17,18,19]
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