Biology subjects

Bryce-Smith, S.

Publications and source records attributed to Bryce-Smith, S..

5 recordsLinked to original sources

bioRxiv2024

TDP-43 loss induces extensive cryptic polyadenylation in ALS/FTD

Nuclear depletion and cytoplasmic aggregation of the RNA-binding protein TDP-43 is the hallmark of ALS, occurring in over 97% of cases. A key consequence of TDP-43 nuclear loss is the de-repression of cryptic exons. Whilst TDP-43 regulated cryptic splicing is increasingly well catalogued, cryptic alternative polyadenylation (APA) events, which define the 3 end of last exons, have been largely overlooked, especially when not associated with novel upstream splice junctions. We developed a novel bioinformatic approach to reliably identify distinct APA event types: alternative last exons (ALE), 3UTR extensions (3Ext) and intronic polyadenylation (IPA) events. We identified novel neuronal cryptic APA sites induced by TDP-43 loss of function by systematically applying our pipeline to a compendium of publicly available and in house datasets. We find that TDP-43 binding sites and target motifs are enriched at these cryptic events and that TDP-43 can have both repressive and enhancing action on APA. Importantly, all categories of cryptic APA can also be identified in ALS and FTD post mortem brain regions with TDP-43 proteinopathy underlining their potential disease relevance. RNA-seq and Ribo-seq analyses indicate that distinct cryptic APA categories have different downstream effects on transcript and translation. Intriguingly, cryptic 3Exts occur in multiple transcription factors, such as ELK1, SIX3, and TLX1, and lead to an increase in wild-type protein levels and function. Finally, we show that an increase in RNA stability leading to a higher cytoplasmic localisation underlies these observations. In summary, we demonstrate that TDP-43 nuclear depletion induces a novel category of cryptic RNA processing events and we expand the palette of TDP-43 loss consequences by showing this can also lead to an increase in normal protein translation.

neuroscience↗

bioRxiv2024

Widespread 3' UTR splicing regulates expression of oncogene transcripts in sequence-dependent and independent manners

BackgroundSplicing in 3 untranslated regions (3 UTRs) is generally considered a signal to elicit transcript degradation via nonsense-mediated decay (NMD) due to the presence of an exon junction complex (EJC) downstream of the stop codon. However, 3 UTR intron (3UI)-containing transcripts are widespread and highly expressed in both normal tissues and cancers. ResultsHere we present and characterise a novel transcriptome assembly built from 7897 solid tumour and normal samples from The Cancer Genome Atlas. We identify thousands of 3UI-containing transcript isoforms, many of which are expressed across multiple cancer types. We find that the expression of core NMD component UPF1 negatively correlates with global 3UI splicing between normal samples, however this correlation is lost in colon cancer. We find that 3UIs found exclusively within 3 UTRs (bona-fide 3UIs) are not predominantly NMD-sensitising, unlike introns present in 3 UTRs due to premature termination. We identify HRAS as an example where 3UI splicing rescues the transcript from NMD. Bona-fide, but not premature termination codon (PTC) carrying 3UI-transcripts are spliced more in cancer samples compared to matched normals in the majority of cancer types analysed. In colon cancer, differentially spliced 3UI-containing transcripts are enriched in the canonical Wnt signalling pathway, with CTNNB1 being the most over-spliced in colon cancer compared to normal. We show that manipulating Wnt signalling can further regulate splicing of Wnt component transcript 3 UTRs. ConclusionsOur results indicate that 3 UTR splicing is not a rare occurrence, especially in colon cancer, where 3 UTR splicing regulates transcript expression in EJC-dependent and independent manners.

genomics↗

bioRxiv2023

BDNF controls phosphorylation and transcriptional networks governing cytoskeleton organization and axonal regeneration

The cell-intrinsic capacity of neurons to regenerate axons requires widespread coordination of the transcriptome, activation of multiple kinases, and reorganization of the cytoskeleton. Axonal repair is also influenced by extrinsic activating factors, such as neurotrophins. Here, we reveal that brain-derived neurotrophic factor (BDNF) amplifies multiple neuron-intrinsic programs to foster axonal regeneration in human motor neurons. Through metabolic RNA sequencing and phosphoproteomic profiling, we elucidate BDNF signalling and its role in axonal regeneration. We discover that BDNF controls RNA stability and transcriptional programs that converge with regeneration-associated gene (RAG) sets. We further unveil that BDNF governs the phosphorylation of multiple proteins essential for cytoskeletal dynamics, a major determinant of effective nerve regeneration. Using compartmentalized neuronal cultures, we demonstrate that the regeneration driven by BDNF depends on the axon-specific activation of ERK/RSK/S6K kinase pathway. We propose a model in which BDNF augments neuron-intrinsic pathways to drive axonal regeneration in human motor neurons. TeaserBDNF aids nerve repair by fine-tuning the metabolism of RNA and by changing the building blocks of the nerve cell cytoskeleton.

molecular biology↗

bioRxiv2023

Extensible benchmarking of methods that identify and quantify polyadenylation sites from RNA-seq data

The tremendous rate with which data is generated and analysis methods emerge makes it increasingly difficult to keep track of their domain of applicability, assumptions, and limitations and consequently, of the efficacy and precision with which they solve specific tasks. Therefore, there is an increasing need for benchmarks, and for the provision of infrastructure for continuous method evaluation. APAeval is an international community effort, organized by the RNA Society in 2021, to benchmark tools for the identification and quantification of the usage of alternative polyadenylation (APA) sites from short-read, bulk RNA-sequencing (RNA-seq) data. Here, we reviewed 17 tools and benchmarked eight on their ability to perform APA identification and quantification, using a comprehensive set of RNA-seq experiments comprising real, synthetic, and matched 3'-end sequencing data. To support continuous benchmarking, we have incorporated the results into the OpenEBench online platform, which allows for seamless extension of the set of methods, metrics, and challenges. We envisage that our analyses will assist researchers in selecting the appropriate tools for their studies. Furthermore, the containers and reproducible workflows generated in the course of this project can be seamlessly deployed and extended in the future to evaluate new methods or datasets.

bioinformatics↗

bioRxiv2021

Common ALS/FTD risk variants in UNC13A exacerbate its cryptic splicing and loss upon TDP-43 mislocalization

Variants within the UNC13A gene have long been known to increase risk of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), two related neurodegenerative diseases defined by mislocalization of the RNA-binding protein TDP-43. Here, we show that TDP-43 depletion induces robust inclusion of a cryptic exon (CE) within UNC13A, a critical synaptic gene, resulting in nonsense-mediated decay and protein loss. Strikingly, two common polymorphisms strongly associated with ALS/FTD risk directly alter TDP-43 binding within the CE or downstream intron, increasing CE inclusion in cultured cells and in patient brains. Our findings, which are the first to demonstrate a genetic link specifically between loss of TDP-43 nuclear function and disease, reveal both the mechanism by which UNC13A variants exacerbate the effects of decreased nuclear TDP-43 function, and provide a promising therapeutic target for TDP-43 proteinopathies. One-Sentence SummaryShared ALS/FTD risk variants increase the sensitivity of a cryptic exon in the synaptic gene UNC13A to TDP-43 depletion.

neuroscience↗