Large-scale phage-based screening reveals extensive pan-viral mimicry of host short linear motifs

Abstract

Viruses mimic host short linear motifs (SLiMs) to hijack and deregulate cellular functions. Studies of motif-mediated interactions therefore provide insight into virus-host dependencies, and reveal targets for therapeutic intervention. Here, we describe the pan-viral discovery of 1,712 SLiM-based virus-host interactions using a phage peptidome tiling the intrinsically disordered protein regions of 229 RNA viruses. We find mimicry of host SLiMs to be a ubiquitous viral strategy, reveal novel host proteins hijacked by viruses, and identify cellular pathways frequently deregulated by viral motif mimicry. Using structural and biophysical analyses, we show that viral mimicry-based interactions have similar binding strength and bound conformations as endogenous interactions. Finally, we establish polyadenylate-binding protein 1 as a potential target for broad-spectrum antiviral agent development. Our platform enables rapid discovery of mechanisms of viral interference and the identification of potential therapeutic targets which can aid in combating future epidemics and pandemics.

Evaluation of affinity-purification coupled to mass spectrometry approaches for capture of short linear motif-based interactions

“…The G-factor was adjusted using wells only containing the FITC-labeled peptide so that the background fluorescence polarization value would be between 10-40 mP. The affinity between the FITC-labeled peptides and the purified TSG101 has been determined previously using saturation binding experiments (17). To determine the affinities of the unlabeled peptides the displacement experiments were performed, where the constant concentration of FITC-labeled peptide (10 nM) and TSG101 UEV (8 μM) was challenged with increasing concentration of unlabeled displacer peptide (1:1 dilution series with the highest concentration of the peptides being 300 μM).…”

“…However, the last decade has seen development of experimental (11)(12)(13) and computational (14,15) approaches for proteomewide screening of SLiM-based interactions and motif. Proteomic peptide phage display (ProP-PD) has been developed as an efficient approach for proteome-wide (and even pan-viral) screening of SLiM-based interactions (7,16,17). ProP-PD screening provides large-scale data on motif-based interactions from selections against a set of bait proteins.…”

Identification of motif-based interactions between SARS-CoV-2 protein domains and human peptide ligands pinpoint antiviral targets

“…Fluorescence polarization experiments were performed as previously described in detail 7 . Briefly, peptides were ordered either unlabeled or as FITC-labelled constructs from GeneCust.…”

“…While most drugs work by inhibiting enzymes, the pharmaceutical industry is turning its attention to the more challenging but largely untouched area of protein-protein interaction interfaces. To this end, several approaches have been developed to identify novel drug targets by mapping the virus-host protein-protein interactome [4][5][6][7] . In a particular class of protein-protein interactions, an intrinsically disordered region (IDR) of a protein interacts with a folded domain of the binding partner.…”

“…Consequently, SLiMs can evolve ex nihilo through accumulation of a few mutations. Because viruses evolve relatively rapidly, viral mimicry of host protein SLiMs has been proposed as a commonly used strategy, and several examples have been described across most viral clades [6][7][8] . Proteins expressed from viral genomes therefore often contain SLiMs which compete with native host protein-protein interactions to hijacking cellular machinery.…”