Replication and transcription machinery for ranaviruses: components, correlation, and functional architecture

Area of Science:

• Virology
• Molecular Biology
• Genomics

Background:

• Ranaviruses are promiscuous pathogens infecting diverse poikilotherm species.
• Replication and transcription mechanisms of these nucleocytoplasmic large DNA viruses (NCLDVs) are largely unknown.
• This study focuses on Andrias davidianus ranavirus (ADRV) and Rana grylio virus (RGV).

Purpose of the Study:

• To screen and uncover the replication and transcription machinery of ADRV and RGV.
• To elucidate the protein components and interactions involved in viral replication and transcription.
• To construct an architectural model of the ranavirus replication and transcription apparatus.

Main Methods:

• Isolation of proteins on nascent DNA.
• Recombinant virus-based affinity purification.
• NanoLuc complementation assay for protein interactions.

Main Results:

• Identified a complex apparatus with at least 30 viral and 6 host proteins.
• Core viral DNA replication proteins (vDPOL, vPCNA, vSSB, vhelicase/primase) were identified.
• Viral RNA polymerase incorporates host RNAPII subunits (Rpb3, Rpb6, Rpb11), a novel finding in NCLDVs.
• Viral DNA polymerase (vDPOL) acts as a central regulatory factor, interacting with multiple viral and host proteins.

Conclusions:

• A comprehensive architecture for ranavirus replication and transcription was established, involving up to 36 components.
• Viral DNA polymerase (vDPOL) is a key central factor coordinating replication and transcription.
• Findings provide a cornerstone for understanding ranavirus replication, progeny production, and cross-species adaptation.