Respiratory syncytial virus ribonucleoproteins hijack microtubule Rab11 dependent transport for intracellular trafficking

PLoS Pathog. 2022 Jul 7;18(7):e1010619. doi: 10.1371/journal.ppat.1010619. eCollection 2022 Jul.

Abstract

Respiratory syncytial virus (RSV) is the primary cause of severe respiratory infection in infants worldwide. Replication of RSV genomic RNA occurs in cytoplasmic inclusions generating viral ribonucleoprotein complexes (vRNPs). vRNPs then reach assembly and budding sites at the plasma membrane. However, mechanisms ensuring vRNPs transportation are unknown. We generated a recombinant RSV harboring fluorescent RNPs allowing us to visualize moving vRNPs in living infected cells and developed an automated imaging pipeline to characterize the movements of vRNPs at a high throughput. Automatic tracking of vRNPs revealed that around 10% of the RNPs exhibit fast and directed motion compatible with transport along the microtubules. Visualization of vRNPs moving along labeled microtubules and restriction of their movements by microtubule depolymerization further support microtubules involvement in vRNPs trafficking. Approximately 30% of vRNPs colocalize with Rab11a protein, a marker of the endosome recycling (ER) pathway and we observed vRNPs and Rab11-labeled vesicles moving together. Transient inhibition of Rab11a expression significantly reduces vRNPs movements demonstrating Rab11 involvement in RNPs trafficking. Finally, Rab11a is specifically immunoprecipitated with vRNPs in infected cells suggesting an interaction between Rab11 and the vRNPs. Altogether, our results strongly suggest that RSV RNPs move on microtubules by hijacking the ER pathway.

MeSH terms

  • Endosomes / metabolism
  • Humans
  • Microtubules / metabolism
  • Protein Transport / physiology
  • Respiratory Syncytial Virus, Human* / metabolism
  • Ribonucleoproteins* / metabolism
  • Viral Proteins / metabolism
  • rab GTP-Binding Proteins* / metabolism

Substances

  • Ribonucleoproteins
  • Viral Proteins
  • rab11 protein
  • rab GTP-Binding Proteins

Grants and funding

MARW was supported by ATIP-AVENIR INSERM program (2018)(https://www.inserm.fr/nous-connaitre/programme-atip-avenir/), and the Fondation Del Duca - Institut de France (https://www.fondation-del-duca.fr/). GC and KM doctoral scholarships were supported by the Versailles St. Quentin university. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.