Rxivist combines preprints from bioRxiv with data from Twitter to help you find the papers being discussed in your field. Currently indexing 57,294 bioRxiv papers from 263,837 authors.
Flaviviruses such as dengue (DENV), Zika (ZIKV) or West Nile virus (WNV) translate their genome as a single multi-pass transmembrane (TM) protein at the endoplasmic reticulum (ER) membrane. Several genetic knockout (KO) screens identified the ER membrane protein complex (EMC) as a critical host component for flavivirus infection. The EMC facilitates accurate insertion, topology and/or stabilization of specific cellular TM proteins including a subset of tail-anchored proteins and G protein-coupled receptors. Here, we show that deletion of EMC in human cell lines decreased infection with DENV, ZIKV and WNV by 100 to 10,000-fold. Using replicon and immunoblotting studies in EMC KO cells, we demonstrated that the EMC was essential for viral protein expression. Ribosome Profiling of DENV-infected wild-type and EMC KO cells revealed no drastic differences in translation efficiency. Instead, absence of EMC led to a large fraction of expressed viral proteins being targeted to the proteasome post-translationally. We detected a decrease in stability in NS4A-NS4B of the viral polyprotein, a region rich in transmembrane domains. Additionally, we identified non-synonymous point mutations in NS4A and NS4B by performing iterative passaging of DENV on EMC KO cells. Adaptive mutations rescued both viral replication and stable expression of the NS4A-NS4B polyprotein segment in EMC KO cells. Lastly, we showed a physical interaction between the EMC and DENV NS4B protein post-cleavage and rapid degradation of processed NS4B in the absence of EMC. Together, our results suggest that the EMC engages with DENV polyproteins to ensure proper biogenesis of the NS4A-NS4B region, and provide further evidence for the cellular function of the EMC in the stable expression of TM proteins.
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