MURV-seq reveals the genome-wide diversity of 5’ termini and leader sequences across poxvirus transcripts

ABSTRACT: Viral RNA polymerases (vRNAPs) can slip on homopolymeric sequences to expand their coding potential in diverse ways. This includes poxviruses, whose vRNAP slips at thymidine-rich motifs to generate non-templated polyadenosine (polyA) leaders on certain transcripts. However, we lack a comprehensive genome-wide understanding of poxviral 5’ leaders and their capping status. Here, we developed Mapping 5′ Untranslated Regions of Viral transcripts (MURV-seq), a customized transcript capture, sequencing and data analysis strategy that distinguishes 5′ triphosphate (5′ppp) versus 7-methylguanosine (m7G) termini and provides high-resolution sequences for the leaders of all annotated poxvirus open reading frames (ORFs). MURV-seq reveals extensive diversity and temporal shifts in the use of templated and non-templated leaders by individual genes. We further define three distinct polyA leader types and refine our understanding of variance in their lengths both between kinetic classes and across individual genes. Contrary to recent suggestions that long polyA leaders generally lack canonical caps, we find that this only occurs on a small fraction of transcripts while all viral transcript types predominantly retain m7G caps. Moreover, while many early genes utilize templated leaders, as expected, transcription upstream of canonical vRNAP slippage sites produces alternative templated leaders for many intermediate and late ORFs. Computational analysis suggests that some of these harbor short upstream ORFs that likely limit their use as 5’ untranslated regions (UTRs), while others are predicted to be functional for the primary ORF or in producing N-terminally extended isoforms. Combined, MURV-seq provides a genome-wide framework for understanding poxvirus transcription start sites and leader diversity.

ORGANISM(S): Homo sapiens

PROVIDER: GSE329296 | GEO | 2026/04/30

REPOSITORIES: GEO