Biology subjects

Aspden, J.

Publications and source records attributed to Aspden, J..

9 recordsLinked to original sources

bioRxiv2026

Constructive neutral evolution explains the emergence of specialised ribosomes in diverse eukaryotes.

Throughout eukaryotic evolution, the structure of the ribosome has been highly conserved, featuring 80 common protein gene families. However, in many eukaryotes, paralogs of these proteins are present. "Specialised ribosomes" have been documented across diverse groups of eukaryotes where they play an important role in the regulation of translation of specific mRNAs. In the case of specialised ribosomes it has been documented that assembled ribosomes that contain specific paralogs can directly affect translational output. This has been proposed to contribute to the regulation of complex responses to environmental change and to coordinate cell-type specific physiology. This poses the question of whether ribosome specialisation principally emerges under an adaptive or neutral model of evolution. Using gene tree-species tree reconciliation, we test competing hypotheses regarding the evolutionary drivers of ribosome specialisation. We determine that examples of specialisation tend to emerge by independent duplication of the same ribosomal proteins in different lineages. We show that pathways to specialisation through paralog formation have arisen independent of: (i) paralog location within the 3D ribosome complex, and (ii) positive selection in these paralogs. We determine that the generalisable model of best fit for the evolution of paralog-mediated eukaryotic ribosomal specialisation is one of constructive neutral evolution. In lineages with small effective population sizes and increased complexity, the emergence and retention of ribosomal protein paralogs has provided the raw material for ratcheting and the emergence of translational regulation at the level of the ribosome.

evolutionary biology↗

bioRxiv2025

Characterising the effect of age and sex on post-transcriptional regulation in synovial joint tissues

Age and sex are major risk factors for joint degeneration and disease susceptibility. While epigenetic and post-transcriptional mechanisms are known to be influenced by these factors, their effects on mRNA kinetics and configuration in musculoskeletal tissues remain poorly defined. To address this gap, we used an equine model to examine how age and sex impact mRNA stability in joint tissues. We measured global transcript half-life in primary chondrocytes from young and old female and young male horses using SLAM-seq. Polyadenylation patterns were additionally analysed in cartilage and synovium across age groups. Our findings demonstrated that age and sex exert distinct regulatory influences on post-transcriptional gene expression in joint tissues. Specifically, ageing alters polyadenylation site usage and transcript turnover in cartilage, even in absence of overt pathology, suggesting that molecular ageing may precede and predispose to joint degeneration. We also found that mean RNA half-life differed significantly between young females and males, with male chondrocytes showing greater transcript stability. This difference suggests that sex-specific regulatory mechanisms may influence RNA stability, which could contribute to differential susceptibility to joint degeneration. Together, these results point to age and sex as key drivers of post-transcriptional regulation with potential roles in shaping joint health trajectories. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=79 SRC="FIGDIR/small/677035v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@99310borg.highwire.dtl.DTLVardef@162ee69org.highwire.dtl.DTLVardef@125108org.highwire.dtl.DTLVardef@860828_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2024

EMG1 methyltransferase activity regulates ribosome occupancy at viral uORFs

Viruses do not encode their own translational machinery and rely exclusively on the translational machinery of the host cell for the synthesis of viral proteins. Viruses have evolved diverse mechanisms to redirect the cellular translation apparatus to favour viral transcripts. A unique mechanism employed by Kaposis sarcoma-associated herpesvirus (KSHV) involves manipulation of cellular ribosome composition, producing virus-induced specialised ribosomes. These ribosomes scan through KSHV uORFs in late lytic genes, allowing efficient translation of downstream main KSHV ORFs. Herein, we highlight the enhanced association of the ribosomal biogenesis factor EMG1 with precursor-40S ribosome complexes during KSHV lytic replication. Depletion of EMG1 results in significantly reduced expression of viral proteins and progression through the lytic cascade, culminating in a dramatic reduction of infectious virus production. We further demonstrate that the methyltransferase activity of EMG1 is required for effective regulation of translation of KSHV uORFs in late lytic genes.

microbiology↗

bioRxiv2023

Differences in binding preferences for XIST partners are observed in mammals with different early pregnancy morphologies

In comparison to protein-coding genes, long non-coding RNAs (lncRNAs) are poorly conserved between species at the sequence level yet they often perform conserved and essential regulatory roles. The lncRNA XIST mediates X chromosome inactivation (XCI) through the interaction of numerous proteins, most of which bind to its repeat regions. Whilst XIST is present across placental mammals, its sequence is divergent. In addition, the timing and mechanistic details of the process of XCI also vary across placental mammals. Here, we sought to determine whether XIST interactor proteins previously identified in a model mammalian species (mouse) also bind in other mammals which exhibit divergent timing of XCI (human and bovine) and differing pre-implantation embryo morphology. We show that XIST and its putative protein partners are coordinately expressed in the endometria of mammals with different pre-implantation embryo morphologies. RNA immunoprecipitations revealed that human WTAP, SPEN, hnRNPK and CIZ1 proteins bind human XIST. CIZ1-XIST binding was also detected in bovine. In both human and cow, we found CIZ1 binds to the repeat E element of XIST. However, human CIZ1 protein is not able to bind bovine repeat E, indicating a species-specific binding interaction. Together these data indicate that several of the key RNA-protein interactions with XIST are common across placental mammals, even though we observe divergence in both the RNA sequence of XIST, and early embryo morphologies. This work sheds light upon the evolution of RNA-protein binding interactions, revealing that the binding events can be conserved even when the precise mechanism of binding has changed. Statements and DeclarationsAuthors have no financial or non-financial interests that are directly or indirectly related to the work submitted for publication.

developmental biology↗

bioRxiv2023

m6A regulates the stability of cellular transcripts required for efficient KSHV lytic replication

The epitranscriptomic modification N6-methyladenosine (m6A) is a ubiquitous feature of the mammalian transcriptome. It modulates mRNA fate and dynamics to exert regulatory control over numerous cellular processes and disease pathways, including viral infection. Kaposis sarcoma-associated herpesvirus (KSHV) reactivation from the latent phase leads to redistribution of m6A topology upon both viral and cellular mRNAs within infected cells. Here we investigate the role of m6A in cellular transcripts upregulated during KSHV lytic replication. Results show that m6A is crucial for the stability of the GPRC5A mRNA, whose expression is induced by the KSHV latent-lytic switch master regulator, the replication and transcription activator (RTA) protein. Moreover, we demonstrate that GPRC5A is essential for efficient KSHV lytic replication by directly regulating NF{kappa}B signalling. Overall, this work highlights the central importance of m6A in modulating cellular gene expression to influence viral infection. Author SummaryChemical modifications on mRNA, such as m6A, are functionally linked to all stages of mRNA metabolism and regulate a variety of biological processes. As such, m6A modification offers unique possibilities for viruses to modulate both viral and host gene expression. m6A has been identified on transcripts encoded by a wide range of viruses and studies to investigate m6A function have highlighted distinct roles in virus life cycles. In addition, cellular transcripts undergoing differential m6A status during infection may also be important for virus replication. In this study we investigate the impact of differential m6A modification in host transcripts during KSHV lytic replication, by identifying transcripts with altered methylation profiles between latent and lytic replication programmes. We show that increased m6A content in one of these cellular mRNAs, GPRC5A, enhances its stability and correlates with increased abundance during KSHV lytic replication. Moreover, the importance of GPRC5A is demonstrated by depletion studies, showing that GPRC5A enhances KSHV lytic replication by inhibiting cell signalling pathways.

microbiology↗

bioRxiv2022

Identification of cancer drivers from tumor-only RNA-seq with RNA-VACAY

Detecting somatic mutations is a cornerstone of cancer genomics and clinical genotyping; however, there has been little systematic evaluation of the utility of RNA sequencing (RNA-seq) for somatic variant detection and driver mutation analysis. Variants found in RNA-Seq are also expressed, reducing the identification of passenger mutations and would not suffer from annotation bias observed in whole-exome sequencing (WES). We developed RNA-VACAY, a containerized pipeline that automates somatic variant calling from tumor RNA-seq data, alone, and evaluated its performance on simulated data and 1,349 RNA-seq samples with matched whole-genome sequencing (WGS). RNA-VACAY was able to detect at least 1 putative driver gene in 15 out of 16 cancer types and identified known driver mutations in 5 and 3 UTRs. The computational cost and time to generate and analyze RNA-seq data is lower than WGS or WES, which decreases the resources necessary for somatic variant detection. This study demonstrates the utility of RNA-seq to detect cancer drivers.

genomics↗

bioRxiv2022

Kaposi's sarcoma-associated herpesvirus induces specialised ribosomes to efficiently translate viral lytic mRNAs

Historically, ribosomes have been viewed as unchanged homogeneous macromolecular machines with no intrinsic regulatory capacity for mRNA translation. However, an emerging concept is that heterogeneity of ribosomal composition exists, which can exert a regulatory function or specificity in translational control. This is supported by recent discoveries identifying compositionally distinct specialised ribosomes that actively regulate mRNA translation. Viruses lack their own translational machinery and impose a high translational demand on the host cell during replication. Here we explore the possibility that Kaposis sarcoma-associated herpesvirus (KSHV) can manipulate host ribosome biogenesis during infection to produce specialised ribosomes which preferentially translate viral transcripts. Quantitative proteomic analysis has identified changes in the stoichiometry and composition of precursor ribosomal complexes during the switch from latent to lytic KSHV replication. Intriguingly, we demonstrate the enhanced association of ribosomal biogenesis factors BUD23 and NOC4L, and a previously uncharacterised KSHV lytic protein, ORF11, with small ribosomal subunit precursor complexes during lytic KSHV infection. Notably, BUD23 depletion resulted in significantly reduced viral gene expression and progression through the lytic cascade, culminating in a dramatic reduction of infectious virion production. Importantly, ribosome profiling demonstrated that BUD23 is essential for the reduced association of ribosomes with KSHV uORFs in late lytic genes, required for the efficient translation of the main open reading frame. Together our results provide new mechanistic insights into KSHV-mediated manipulation of cellular ribosome composition inducing a population of specialised ribosomes to facilitate efficient translation of viral mRNAs.

microbiology↗

bioRxiv2020

Ribosome fingerprinting with a solid-state nanopore

Nanopores hold great potential for the analysis of complex biological molecules at the single entity level. One particularly interesting macromolecular machine is the ribosome, responsible for translating mRNA into proteins. In this study, we use a solid-state nanopore to fingerprint 80S ribosomes and polysomes from a human neuronal cell line and, Drosophila melanogaster cultured cells and ovaries. Specifically, we show that the peak amplitude and dwell time characteristics of 80S ribosomes are distinct from polysomes and can be used to discriminate ribosomes from polysomes in mixed samples. Moreover, we are able to distinguish large polysomes, containing more than 7 ribosomes, from those containing 2-3 ribosomes, and demonstrate a correlation between polysome size and peak amplitude. This study highlights the application of solid-state nanopores as a rapid analytical tool for the detection and characterization of ribosomal complexes.

biophysics↗

bioRxiv2019

Developmental regulation of Canonical and small ORF translation from mRNA

Ribosomal profiling has revealed the translation of thousands of sequences outside of annotated protein-coding genes, including small Open Reading Frames of less than 100 codons, and the translational regulation of many genes. Here we have improved Poly-Ribo-Seq and applied it to Drosophila melanogaster embryos to extend the catalogue of in-vivo translated small ORFs, and to reveal the translational regulation of both small and canonical ORFs from mRNAs across embryogenesis. We obtain highly correlated samples across five embryonic stages, with close to 500 million putative ribosomal footprints mapped to mRNAs, and compared them to existing Ribo-Seq and proteomic data. Our analysis reveals, for the first time in Drosophila, footprints mapping to codons in a phased pattern, the hallmark of productive translation, and we propose a simple binomial probability metric to ascertain translation probability. However, our results also reveal reproducible ribosomal binding apparently not resulting in productive translation. This non-productive ribosomal binding seems to be especially prevalent amongst upstream short ORFs located in the 5 mRNA Leaders, and amongst canonical ORFs during the activation of the zygotic translatome at the maternal to zygotic transition. We suggest that this non-productive ribosomal binding might be due to cis-regulatory ribosomal binding, and to defective ribosomal scanning of ORFs outside periods of productive translation. Finally, we show that the main function of upstream short ORFs is to buffer the translation of canonical ORFs, and that in general small ORFs in mRNAs display Poly-Ribo-Seq and bioinformatics markers compatible with an evolutionary transitory state towards full coding function.

genomics↗