Abstract
Genetic perturbation screens using RNA interference (RNAi) have been conducted successfully to identify host factors that are essential for the life cycle of bacteria or viruses. So far, most published studies identified host factors primarily for single pathogens. Furthermore, often only a small subset of genes, e.g., genes encoding kinases, have been targeted. Identification of host factors on a pan-pathogen level, i.e., genes that are crucial for the replication of a diverse group of pathogens has received relatively little attention, despite the fact that such common host factors would be highly relevant, for instance, for devising broad-spectrum anti-pathogenic drugs. Here, we present a novel two-stage procedure for the identification of host factors involved in the replication of different viruses using a combination of random effects models and Markov random walks on a functional interaction network. We first infer candidate genes by jointly analyzing multiple perturbations screens while at the same time adjusting for high variance inherent in these screens. Subsequently the inferred estimates are spread across a network of functional interactions thereby allowing for the analysis of missing genes in the biological studies, smoothing the effect sizes of previously found host factors, and considering a priori pathway information defined over edges of the network. We applied the procedure to RNAi screening data of four different positive-sense single-stranded RNA viruses, Hepatitis C virus, Chikungunya virus, Dengue virus and Severe acute respiratory syndrome coronavirus, and detected novel host factors, including UBC, PLCG1, and DYRK1B, which are predicted to significantly impact the replication cycles of these viruses. We validated the detected host factors experimentally using pharmacological inhibition and an additional siRNA screen and found that some of the predicted host factors indeed influence the replication of these pathogens.
Highlights
Genetic perturbation screens, such as RNA interference (RNAi) and CRIPSR-Cas9 screens, allow for the detection of host dependency and restriction factors by perturbing a target gene or transcript and observing its impact on the life cycle of a pathogen
The identification of host factors that are essential for viral replication is of scientific interest, and of clinical relevance, since they could serve as targets for the development of antiviral therapies, which is still unavailable for many important pathogens
In the case of positive-sense single-stranded RNA (ssRNA) viruses a variety of such host factors have been earlier described and experimentally verified
Summary
Genetic perturbation screens, such as RNA interference (RNAi) and CRIPSR-Cas screens, allow for the detection of host dependency and restriction factors by perturbing a target gene or transcript and observing its impact on the life cycle of a pathogen. In RNAi screens, genes are perturbed with small interferring RNAs (siRNAs). For virtually all of these steps, the virus strongly depends on host proteins due to the small RNA virus genomes with limited coding capacity. Another common feature of +RNA viruses is that their RNA synthesis takes place in specialized structures that are associated with modified host membranes [10]. In order to understand the virus-host interplay reliable identification of potential host factors involved in virus replication is crucial
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