Biology subjects

Bayfield, M. A.

Publications and source records attributed to Bayfield, M. A..

7 recordsLinked to original sources

bioRxiv2026

Endonuclease G governs an antiviral RNA surveillance system in budding yeast

Endonuclease G family DNA/RNA nucleases localize to the mitochondrial intermembrane space and prevent escape of mitochondrial DNA to the cytosol or nucleus. While extramitochondrial Endonuclease G also targets nuclear DNA during cell death, RNA regulatory roles of this family remain enigmatic. Here we discover an RNA surveillance system in Saccharomyces cerevisiae governed by Nuc1, the yeast Endonuclease G homolog. This system limits replication of L-A, an RNA mycovirus that endemically infects this organism. Whereas L-A resides in the cytosol, we find that Nuc1's antiviral function requires its mitochondrial intermembrane space localization. To understand Nuc1's indirect antiviral mechanism we investigated its genetic interactions with REX1 and REX2, RNA exonucleases that control 3' end processing of tRNA and rRNA intermediates in the nucleus. We demonstrate that the nuc1{triangleup} antiviral defect is massively amplified by rex1{triangleup} rex2{triangleup} leading to L-A RNA levels that surpass mature rRNAs and overwhelm the cells. NUC1 concurrently controls a pool of mis-processed tRNA and rRNA species that accumulate in rex1{triangleup} rex2{triangleup} mutants. Linking these mis-processed RNAs with L-A control, we show they are significantly reduced in L-A infected cells, suggesting that L-A depletes them to enhance its replication. Our findings identify a previously unrecognized RNA surveillance function of Nuc1 that prevents aberrant rRNA and tRNA species from promoting replication of the L-A mycovirus.

molecular biology↗

bioRxiv2025

SnoBIRD: A tool to identify C/D box snoRNAs and refine their annotation across all eukaryotes

Small nucleolar RNAs (snoRNAs), a group of noncoding RNAs present amongst all eukaryotes, are most extensively characterized for their regulation of ribosome biogenesis and splicing. Despite their central roles, current snoRNA annotations remain incomplete. Several eukaryote genome annotations contain few or no snoRNAs, and none distinguish expressed snoRNAs from their pseudogenes--a recently characterized snoRNA subclass with distinct features and expression levels. To address this, we developed SnoBIRD, a BERT-based C/D box snoRNA predictor trained on snoRNAs spanning all eukaryote kingdoms. We show that SnoBIRD outperforms existing tools and is the only predictor capable of identifying snoRNA pseudogenes using biologically relevant signal. Applied on the fission yeast and human genomes, we demonstrate that only SnoBIRD scales well with genome size in terms of runtime, and we identify and experimentally validate several new SnoBIRD-predicted C/D box snoRNAs. By running SnoBIRD on multiple eukaryote genomes, we identify hundreds of novel snoRNA candidates and highlight SnoBIRDs usefulness to determine the evolutionary paths of snoRNAs distributed across different species. Overall, SnoBIRD represents a user-friendly and efficient tool for reliably predicting C/D box snoRNAs and their pseudogenes across any eukaryote genome. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=109 SRC="FIGDIR/small/646650v1_ufig1.gif" ALT="Figure 1"> View larger version (19K): org.highwire.dtl.DTLVardef@99b390org.highwire.dtl.DTLVardef@dbe375org.highwire.dtl.DTLVardef@32358aorg.highwire.dtl.DTLVardef@58fb3a_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

bioRxiv2024

Respiratory Syncytial Virus (RSV) optimizes the translational landscape during infection

Viral infection often triggers eukaryotic initiator factor 2 (eIF2) phosphorylation, leading to global 5-cap-dependent translation inhibition. RSV encodes messenger RNAs (mRNAs) mimicking 5-cap structures of host mRNAs and thus inhibition of cap-dependent translation initiation would likely also reduce viral translation. We confirmed that RSV limits widespread translation initiation inhibition and unexpectedly found that the fraction of ribosomes within polysomes increases during infection, indicating higher ribosome loading on mRNAs during infection. We found that AU-rich host transcripts that are less efficiently translated under normal conditions become more efficient at recruiting ribosomes, similar to RSV transcripts. Viral transcripts are transcribed in cytoplasmic inclusion bodies, where the viral AU-rich binding protein M2-1 has been shown to bind viral transcripts and shuttle them into the cytoplasm. We further demonstrated that M2-1 is found on polysomes, and that M2-1 might deliver host AU-rich transcripts for translation. ImportanceViruses strongly rely on the hosts translational machinery to produce viral proteins required for replication. However, it is unknown how viruses that do not globally inhibit cap-dependent translation compete with abundant host transcripts for ribosomes. In this study, we found that respiratory syncytial virus (RSV) infection results in redistribution of 80S monosomes into the polysomes. High-throughput sequencing of translating transcripts revealed that low translation efficiency transcripts become more efficient at ribosome recruitment which are virus-resembling AU-rich host transcripts. Finally, we also uncover that AU-rich RNA binding protein RSV-M2-1 interacts with polysomes through contacts to mRNA. These findings revealed that RSV optimizes the translational landscape rather than inhibiting host translation.

molecular biology↗

bioRxiv2023

The tRNA methyltransferase Trm1 promotes eukaryotic pre-tRNA maturation through catalytic and non-catalytic activities

tRNAs undergo an extensive maturation process involving post-transcriptional modifications often associated with tRNA structural stability and promoting the native fold. Impaired post-transcriptional modification has been linked to human disease, likely through defects in translation, mitochondrial function, and increased susceptibility to degradation by various tRNA decay pathways. More recently, evidence has emerged that bacterial tRNA modification enzymes can act as tRNA chaperones to guide tRNA folding in a manner independent from catalytic activity. Here, we provide evidence that the fission yeast tRNA methyltransferase Trm1, which dimethylates nuclear- and mitochondrial-encoded tRNAs at G26, can also promote tRNA functionality in the absence of catalysis. We show that wild type and catalytic-dead Trm1 are active in an in vivo tRNA-mediated suppression assay and possess RNA strand annealing and dissociation activity in vitro, similar to previously characterized RNA chaperones. Trm1 and the RNA chaperone La have previously been proposed to function synergistically in promoting tRNA maturation, yet we surprisingly demonstrate that La binding to nascent pre-tRNAs decreases Trm1 tRNA dimethylation in vivo and in vitro. Collectively, these results support the hypothesis for tRNA modification enzymes that combine catalytic and non-catalytic activities to promote tRNA maturation, as well as expand our understanding of how La function can influence tRNA modification.

molecular biology↗

bioRxiv2023

The fission yeast methylphosphate capping enzyme Bmc1/Bin3 promotes 2'-O- methylation of U6 and pre-mRNA splicing

Splicing requires the tight coordination of dynamic spliceosomal RNAs and proteins. U6 is the only spliceosomal RNA transcribed by RNA Polymerase III and undergoes an extensive maturation process. In humans and fission yeast, this includes addition of a 5 {gamma}-monomethyl phosphate cap by members of the Bin3/MePCE family. Previously, we have shown that the Bin3/MePCE homolog Bmc1 is recruited to the S. pombe telomerase holoenzyme by the LARP7 family protein Pof8, where it acts in a catalytic-independent manner to protect the telomerase RNA and facilitate holoenzyme assembly. Here, we show that Bmc1 and Pof8 also interact in a U6-containing snRNP. We demonstrate that Bmc1 and Pof8 promote 2-O-methylation of U6 and identify and characterize a non-canonical snoRNA that guides this methylation. Further, we show that fission yeast strains deleted of Bmc1 or Pof8 show altered U6 snRNP assembly patterns, supporting a more general role for these factors in guiding noncoding RNP assembly beyond the telomerase RNP. These results are thus consistent with a novel role for Bmc1/MePCE family members in stimulating U6 post-transcriptional modifications.

molecular biology↗

bioRxiv2022

Altered tRNA processing is linked to a distinct and unusual La protein in Tetrahymena thermophila

Nascent pre-tRNAs are transcribed by RNA polymerase III and immediately bound by La proteins on the UUU-3OH sequence, using a tandem arrangement of the La motif and an adjacent RNA recognition motif-1 (RRM1), resulting in protection from 3-exonucleases and promotion of pre-tRNA folding. The Tetrahymena thermophila protein Mlp1 has been classified as a genuine La protein, despite the predicted absence of the RRM1. We found that Mlp1 functions as a La protein through binding of pre-tRNAs and affecting processing in Tetrahymena thermophila and when expressed in fission yeast. However, unlike in other examined eukaryotes, depletion of Mlp1 results in 3-trailer stabilization. We also observed that 3-trailers in Tetrahymena thermophila are uniquely short relative to other examined eukaryotes, and that 5-leaders have evolved to disfavour pre-tRNA leader/trailer pairing. Our data indicate that this variant Mlp1 architecture is linked to an altered, novel mechanism of tRNA processing in Tetrahymena thermophila.

molecular biology↗

bioRxiv2021

The methyl phosphate capping enzyme Bin3 is a stable component of the fission yeast telomerase holoenzyme

The telomerase holoenzyme is critical for maintaining eukaryotic genome integrity. In addition to a reverse transcriptase and an RNA template, telomerase contains additional proteins that protect the telomerase RNA and promote holoenzyme assembly. Here we report that the methyl phosphate capping enzyme (MePCE) Bin3 is a stable component of the S. pombe telomerase holoenzyme. Bin3 associates with the telomerase holoenzyme and U6 snRNA through an interaction with the recently described LARP7 family member Pof8, and we demonstrate that these two factors are evolutionarily linked in fungi. Our data suggest that the association of Bin3 with telomerase is independent of its methyltransferase activity, but rather that Bin3 functions in telomerase holoenzyme assembly by promoting TER1 accumulation and Pof8 recruitment to TER1. Taken together, this work yields new insight into the composition, assembly, and regulation of the telomerase holoenzyme in fission yeast as well as the breadth of its evolutionary conservation.

molecular biology↗