cis-Acting Element at the 5' Noncoding Region of Tacaribe Virus S RNA Modulates Genome Replication

Area of Science:

• Virology
• Molecular Biology
• RNA Virus Replication

Background:

• Tacaribe virus (TCRV) is a prototype New World mammarenavirus, a group known for causing human hemorrhagic fevers.
• The TCRV genome has small (S) and large (L) RNA segments, each with 5' and 3' noncoding regions (NCRs).
• The function of internal noncoding regions (iNCRs) in TCRV S RNA synthesis is not well understood, despite the known importance of terminal sequences for promoter function.

Purpose of the Study:

• To investigate the role of the 5' and 3' internal noncoding regions (iNCRs) in Tacaribe virus (TCRV) S RNA synthesis.
• To determine if manipulating the TCRV S genomic 5' iNCR sequence can lead to viral attenuation for vaccine development.

Main Methods:

• Generation of mutant S-like minigenomes and miniantigenomes.
• Utilized minireplicon assays, Northern blotting, and reverse transcription-quantitative PCR (RT-qPCR).
• Employed a Tacaribe virus (TCRV) reverse genetic approach for in vivo virulence studies.

Main Results:

• The genomic 5' iNCR is specifically involved in minigenome replication but not transcription.
• The S genome 3' iNCR plays a minimal role in TCRV S RNA synthesis.
• The integrity of the entire genomic 5' iNCR is essential; specific secondary structures or RNA-RNA interactions are not strictly required.
• Mutagenesis of the 5' promoter-proximal subregion slightly reduced recombinant TCRV virulence in vivo.

Conclusions:

• The 5' viral RNA noncoding sequence adjacent to the promoter is critical for Tacaribe virus (TCRV) S RNA replication.
• Findings provide insights into how this 5' noncoding region aids the L polymerase in efficient viral S RNA synthesis.
• Genetic manipulation of New World mammarenavirus sequences, particularly the 5' iNCR, shows potential for designing attenuated recombinant virus vaccine platforms.