Journal Article

Proximity interactome analysis of Lassa polymerase reveals eRF3a/GSPT1 as a druggable target for host-directed antivirals

Pietzsch, Colette, Ting, Alice Y., Cho, Kelvin Frank, Tsaprailis, George, Crynen, Gogce, de la Torre, Juan Carlos, Bukreyev, Alexander, Fang, Jingru, Witwit, Haydar, Saphire, Erica Ollmann

2022

Overview

Completion of the Lassa virus (LASV) life cycle critically depends on the activities of the virally encoded, RNA-dependent RNA polymerase in replication and transcription of the viral RNA genome in the cytoplasm of infected cells. The contribution of cellular proteins to these processes remains unclear. Here, we applied proximity proteomics to define the interactome of LASV polymerase in cells under conditions that recreate LASV RNA synthesis. We engineered a LASV polymerase-biotin ligase (TurboID) fusion protein that retained polymerase activity and successfully biotinylated the proximal proteome, which allowed the identification of 42 high-confidence LASV polymerase interactors. We subsequently performed a small interfering RNA (siRNA) screen to identify those interactors that have functional roles in authentic LASV infection. As proof of principle, we characterized eukaryotic peptide chain release factor subunit 3a (eRF3a/GSPT1), which we found to be a proviral factor that physically associates with LASV polymerase. Targeted degradation of GSPT1 by a small-molecule drug candidate, CC-90009, resulted in strong inhibition of LASV infection in cultured cells. Our work demonstrates the feasibility of using proximity proteomics to illuminate and characterize yet-to-be-defined host-pathogen interactome, which can reveal new biology and uncover novel targets for the development of antivirals against highly pathogenic RNA viruses.

• Publisher: National Academy of Sciences
• Subject: Acetamides - pharmacology / Acetamides - therapeutic use / Antiviral agents / Antiviral Agents - pharmacology / Antiviral Agents - therapeutic use / Biological Sciences / Biotin / Cell Line, Tumor / Chemical synthesis / Cytoplasm / DNA-directed RNA polymerase / Drug development / Fusion protein / Genomes / Humans / Isoindoles - pharmacology / Isoindoles - therapeutic use / Lassa Fever - drug therapy / Lassa virus - drug effects / Life cycles / Microbiology / Peptide Termination Factors - metabolism / Piperidones - metabolism / Piperidones - pharmacology / Piperidones - therapeutic use / Protein Interaction Maps - drug effects / Proteins / Proteolysis - drug effects / Proteome / Proteomes / Proteomics / Proximity / RNA polymerase / RNA viruses / RNA-Dependent RNA Polymerase - metabolism / RNA-directed RNA polymerase / siRNA / Transcription / Viral Proteins - metabolism