Reovirus uses temporospatial compartmentalization to orchestrate core versus outercapsid assembly

Area of Science:

• Virology
• Cell Biology
• Molecular Biology

Background:

• Reoviridae family viruses, like mammalian orthoreovirus (reovirus), replicate by encapsidating double-stranded RNA (dsRNA) within transcriptionally active core capsids.
• Reovirus replication requires amplification of progeny cores and assembly of outer capsid (OC) proteins to form infectious virions.
• Existing models propose simultaneous core amplification and OC assembly in shared viral factories, but this study investigated potential temporal regulation.

Purpose of the Study:

• To investigate the spatiotemporal regulation of reovirus replication, specifically the timing of outer capsid (OC) protein assembly relative to core amplification.
• To elucidate the role of OC proteins, particularly μ1, in the spatial organization and temporal progression of reovirus replication factories.
• To challenge the existing model of shared viral factories and propose an advanced model of reovirus replication.

Main Methods:

• Kinetic analysis of viral RNA and protein production, alongside core and whole virus particle accumulation.
• High-resolution fluorescence and electron microscopy to visualize viral factories and protein localization.
• RNA interference (siRNA) to silence OC μ1 protein expression and assess its impact on factory dynamics.

Main Results:

• Core amplification occurred early at peripheral factories, while OC proteins localized to lipid droplets (LDs) near the nucleus in a μ1-dependent manner.
• OC assembly was temporally delayed relative to core amplification, with viral RNAs and proteins produced simultaneously.
• Silencing μ1 expression resulted in peripheral factories, indicating its role in the transition to perinuclear factories where OC assembly occurred later in infection.
• Reovirus replication involves spatiotemporal segregation of core amplification and OC assembly, with distinct factory locations and timing.

Conclusions:

• Reovirus replication is not confined to shared factories but exhibits spatiotemporal segregation of core amplification and outer capsid assembly.
• Outer capsid protein μ1 plays a critical role in relocating replication factories to the nucleus-proximal region, facilitating OC assembly.
• An advanced model of reovirus replication is proposed, highlighting distinct spatial and temporal dynamics for core amplification and virion maturation.