Biology subjects

Martinez, N. M.

Publications and source records attributed to Martinez, N. M..

3 recordsLinked to original sources

bioRxiv2026

A host enzyme pseudouridylates an RNA bacteriophage genome and accelerates virus production

RNA modifications are prevalent and potent regulators of gene expression across all domains of life. However, whether they exist in bacteriophages and whether they impact phage-host interactions are largely unexplored. To address this, we leveraged the ssRNA bacteriophage MS2 and its E. coli host. Using quantitative nanopore direct RNA sequencing and pseudouridine profiling, we found that essentially all copies of MS2 genomic RNA are pseudouridylated at a single position within the maturation protein gene. Through analysis of the surrounding sequence context, analysis of predicted structure, and genetic assignment of the pseudouridine to its installing enzyme, we demonstrate that the bacterial host enzyme RluA is responsible for pseudouridylation of the phage genome. In the absence of host RluA, the latent and eclipse periods are extended, indicating that the enzyme is important to early stages of phage infectivity. We find that this pseudouridine is present at varying occupancies in additional ssRNA coliphages, also installed by host RluA, and that the conserved features of RluA substrates are present in diverse ssRNA phages with host species beyond E. coli. Altogether, our findings identify RNA modification as a feature of bacteriophage genomes and establish host RNA-modifying enzymes as a class of infection-modulating phage host factors.

molecular biology↗

bioRxiv2025

SWARM: A Single-Molecule Workflow for High-Precision Profiling of RNA Modifications

Nanopore direct RNA sequencing promises to decode the epitranscriptome by detecting multiple modifications on individual RNA molecules, but its potential for biological discovery is hampered by high false-positive rates. We present SWARM, an AI-based framework designed to overcome this fundamental limitation. Its key innovation is a crosstalk-aware training strategy that incorporates non-target modifications and orthogonally validated cellular signals, enabling high-precision detection of m6A, pseudouridine ({Psi}), and m5C at single-nucleotide and single-molecule resolution. Using rigorous in vitro and cellular RNA benchmarks, SWARM outperforms existing tools and maintains strong agreement with orthogonal methods. Applying SWARM across mammalian tissues reveals thousands of novel modification sites with confirmed motifs and localisation patterns. Our high-resolution multi-tissue modification map revealed no evidence of widespread m6A-{Psi} interplay in predominant writer contexts, challenging models of a coordinated epitranscriptomic code. We further discovered a previously unrecognised splicing-shaped mode of {Psi} deposition, whereby TRUB1-mediated pseudouridylation preferentially occurs after exon-exon ligation, consistent with local RNA structure stabilisation. SWARM provides a robust, universally applicable tool for epitranscriptome discovery.

bioinformatics↗

bioRxiv2020

Pseudouridine synthases modify human pre-mRNA co-transcriptionally and affect splicing

Eukaryotic messenger RNAs are extensively decorated with modified nucleotides and the resulting epitranscriptome plays important regulatory roles in cells 1. Pseudouridine ({Psi}) is a modified nucleotide that is prevalent in human mRNAs and can be dynamically regulated 2-5. However, it is unclear when in their life cycle RNAs become pseudouridylated and what the endogenous functions of mRNA pseudouridylation are. To determine if pseudouridine is added co-transcriptionally, we conducted pseudouridine profiling 2 on chromatin-associated RNA to reveal thousands of intronic pseudouridines in nascent pre-mRNA at locations that are significantly associated with alternatively spliced exons, enriched near splice sites, and overlap hundreds of binding sites for regulatory RNA binding proteins. Multiple distinct pseudouridine synthases with tissue-specific expression pseudouridylate pre-mRNA sites, and genetic manipulation of the predominant pre-mRNA modifying pseudouridine synthases PUS1, PUS7 and RPUSD4 induced widespread changes in alternative splicing in cells, supporting a role for pre-mRNA pseudouridylation in alternative splicing regulation. Consistently, we find that individual pseudouridines identified in cells are sufficient to directly affect splicing in vitro. Together with previously observed effects of artificial pseudouridylation on RNA-RNA6-8 and RNA-protein 9-11 interactions that are relevant for splicing, our results demonstrate widespread co-transcriptional pre-mRNA pseudouridylation and establish the enormous potential for this RNA modification to control human gene expression.

molecular biology↗