Preprint Article Version 1 Preserved in Portico This version is not peer-reviewed

Hardening of Respiratory Syncytial Virus Inclusion Bodies by Cyclopamine Proceeds through Perturbation of the Interactions between the M2-1 Protein, RNA and the P Protein

Cédric Diot
,
Charles-Adrien Richard
ORCID logo ,
Jennifer Risso-Ballester
,
Davy Martin
,
Jenna Fix
,
Jean-François Eléouët
ORCID logo ,
Christina Sizun
ORCID logo ,
Marie-Anne Rameix-Welti
* ,
Marie Galloux
* ORCID logo
Version 1 : Received: 25 June 2023 / Approved: 26 June 2023 / Online: 26 June 2023 (09:59:43 CEST)

A peer-reviewed article of this Preprint also exists.

Diot, C.; Richard, C.-A.; Risso-Ballester, J.; Martin, D.; Fix, J.; Eléouët, J.-F.; Sizun, C.; Rameix-Welti, M.-A.; Galloux, M. Hardening of Respiratory Syncytial Virus Inclusion Bodies by Cyclopamine Proceeds through Perturbation of the Interactions of the M2-1 Protein with RNA and the P Protein. Int. J. Mol. Sci. 2023, 24, 13862. Diot, C.; Richard, C.-A.; Risso-Ballester, J.; Martin, D.; Fix, J.; Eléouët, J.-F.; Sizun, C.; Rameix-Welti, M.-A.; Galloux, M. Hardening of Respiratory Syncytial Virus Inclusion Bodies by Cyclopamine Proceeds through Perturbation of the Interactions of the M2-1 Protein with RNA and the P Protein. Int. J. Mol. Sci. 2023, 24, 13862.

Abstract

Respiratory syncytial virus (RSV) RNA synthesis takes place in cytoplasmic viral factories also called inclusion bodies (IBs), which are membrane-less organelles concentrating the viral RNA polymerase complex. IBs assembly is driven by liquid-liquid phase separation promoted by interactions between the viral nucleoprotein N and the phosphoprotein P. We recently demonstrated that cyclopamine (CPM) inhibits RSV multiplication by disorganizing and hardening IBs. Although a single mutation in the viral transcription factor M2-1 induced resistance to CPM, the mechanism of action of CPM still remains to be characterized. Here, using FRAP experiments on reconstituted pseudo-IBs both in cellula and in vitro, we first demonstrated that CPM activity depends on the presence of M2-1 together with N and P. We show that CPM impairs the competition between P and RNA binding to M2-1. As mutations on both P and M2-1 induced resistance against CPM activity, we suggest that CPM may affect the dynamics of the M2-1−P interaction, thereby affecting the relative mobility of proteins contained in RSV IBs. Overall, our results reveal that stabilizing viral protein-protein interactions is an attractive new antiviral approach. They pave the way for the rational chemical optimization of new specific anti-RSV molecules.

Keywords

RSV; cyclopamine; M2-1−P interaction; antiviral mechanism; inclusion bodies

Subject

Biology and Life Sciences, Virology

Copyright: This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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