Extensive High-Order Complexes within SARS-CoV-2 Proteome Revealed by Compartmentalization-Aided Interaction Screening
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Abstract
Bearing the largest single-stranded RNA genome in nature, SARS-CoV-2 utilizes sophisticated replication/transcription complexes (RTCs), mainly composed of a network of nonstructural proteins and nucleocapsid protein, to establish efficient infection. Here, we developed an innovative interaction screening strategy based on phase separation in cellulo , namely co mpartmentalization of p rotein-protein i nteractions in c ells (CoPIC). Utilizing CoPIC screening, we mapped the interaction network among RTC-related viral proteins. We identified a total of 47 binary interactions among 14 proteins governing replication, discontinuous transcription, and translation of coronaviruses. Further exploration via CoPIC led to the discovery of extensive ternary complexes composed of these components, which infer potential higher-order complexes. Taken together, our results present an efficient, and robust interaction screening strategy, and indicate the existence of a complex interaction network among RTC-related factors, thus opening up new opportunities to understand SARS-CoV-2 biology and develop therapeutic interventions for COVID-19.
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SciScore for 10.1101/2020.12.26.424422: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Table 2: Resources
No key resources detected.
Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:There is another caveat associated with CoPIC. If the partner to be recruited can also undergo phase separation, the potentially complicated interaction relationship between the scaffold phase and the client phase might hinder …
SciScore for 10.1101/2020.12.26.424422: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Table 2: Resources
No key resources detected.
Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:There is another caveat associated with CoPIC. If the partner to be recruited can also undergo phase separation, the potentially complicated interaction relationship between the scaffold phase and the client phase might hinder the straightforward recruitment of the clients. For example, for the PPI screening involving the nucleocapsid protein N, there were hardly any positive interactions detected except for its self-association when N protein serves as the client; however multiple PPIs were detected when N is fused with the scaffold, GFP-Nup98N (Figure 3A). We reasoned that the phase separation property of N protein (Iserman et al., 2020; Perdikari et al., 2020; Zhao et al., 2020) interferes with its recruitment into the phase-separated compartment of the scaffolds. To circumvent this caveat, binding partners with phase separation propensity should be fused with the scaffold and those without phase separation propensity should serve as the clients. Even with these caveats considered, CoPIC is a simple and efficient method for investigating PPIs in cells. In addition, CoPIC can be used to detect weak interactions since the signals are enriched within the compartments and the client proteins are overexpressed to high concentrations. Taking the interaction of Nsp14 with Nsp16 as an example, the negative pair demonstrated by the typical co-IP assay turned out to exhibit a salient bidirectional colocalization within the compartment mediated by GFP-Nup98N (Figure 3, S4B, S4C). Not...
Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
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