Preliminary data shows that the alphavirus Semliki Forest virus (SFV) replicates less well in RNase-L knockout (KO) mouse embryonic fibroblasts (MEFs) than in wild-type cells. This is surprising, as RNase-L is generally considered to be an antiviral protein, degrading RNA in response to viral infection (1).
One explanation involves the interaction of alphaviruses with host RNA-binding proteins. Studies performed primarily with the alphavirus Sindbis virus (SINV) have shown the RNA binding protein HuR binds with high affinity to alphavirus RNA through sequence elements in the 3’ untranslated region (UTR), resulting in increased stability of viral transcripts (2, 3). Stability of some host transcripts is also reduced during infection (4). Tristetraprolin (TTP) is another RNA-binding protein that binds competitively with HuR and promotes RNA degradation (5). TTP has been found to recruit RNase L to cellular transcripts under mitogen stimulation, resulting in the degradation of these transcripts (6).
We hypothesise that in SFV infected cells, HuR is sequestered to viral RNA. This protects viral RNA from TTP binding, and therefore degradation by RNase L. Cellular transcripts with no HuR bound would remain vulnerable to TTP binding and RNase L degradation, leading to reduction of cellular transcripts, and reduced competition for translation.
Consistent with studies in SINV, we have shown using immunofluorescent microscopy that HuR relocalises from the nucleus to the cytoplasm during SFV infection. We will determine the roles of HuR, TTP and RNase L in SFV infection using siRNA knockdown of expression of these genes. The effect of RNase L on host transcript levels during SFV infection will be determined using whole-transcriptome sequencing of SFV infected WT and RNase L KO cells.