Genome-wide Viral Nascent RNA Sequencing Unveils Polymerase Pausing Landscape at Single-nucleotide Precision

TenVIP-seq maps non-random RdRp pausing across all eight influenza A virus gene segments at single-nucleotide resolution, revealing distinct pause landscapes on vRNA versus cRNA/mRNA templates.

• Why it matters: Mature-RNA methods miss dynamic transcription/replication intermediates, limiting mechanistic insight needed to translate in vitro findings into host-cell antiviral strategies.
• What they did: The study developed Total Elongating Nascent VIral Polymerase single-molecule Sequencing (TenVIP-seq) to isolate newly synthesized RNA within the RdRp complex and profile genome-wide pausing and terminal misincorporations.
• The result: Favipiravir (T-705) intensified RdRp pauses without adding new sites, TRIM25 knockout reduced pausing globally, and paused-RdRp sequencing detected terminal misincorporations averaging 51.6% (vRNA) and 44.0% (cRNA/mRNA).

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