Transcribing and replicating particles in a double-stranded RNA virus from Leishmania

Area of Science:

• Virology
• Molecular Biology
• Parasitology

Background:

• Double-stranded RNA (dsRNA) viruses replicate through alternating transcription and replication phases.
• RNA-dependent RNA polymerase exhibits both transcriptase and replicase activities.
• Previous studies detected these activities in Leishmania guyanensis viruses in vitro.

Purpose of the Study:

• To elucidate the transcription and replication mechanisms of dsRNA viruses in Leishmania.
• To characterize the polarity of viral RNA products.
• To investigate the relationship between viral replication and host cell density.

Main Methods:

• Northern hybridization to determine RNA polarity.
• Sucrose gradient sedimentation to separate viral particle types.
• In vitro assays to detect viral polymerase activities.

Main Results:

• Single-stranded RNA products were confirmed to be of positive polarity, indicating transcriptase activity.
• Conservative transcription was suggested for Leishmania viruses.
• Two particle types were identified: slower-sedimenting single-stranded RNA particles and faster-sedimenting double-stranded RNA particles.
• Single-stranded RNA particles co-purified with viral replicase activity.
• Replicase activity decreased with increasing parasite density.

Conclusions:

• Leishmania dsRNA viruses likely employ a conservative transcription strategy.
• The findings support a replicative model consistent with other dsRNA viruses.
• Viral replication may be linked to host cell division, as suggested by the inverse correlation with parasite density.