Biology subjects

Barron, N.

Publications and source records attributed to Barron, N..

7 recordsLinked to original sources

bioRxiv2026

Biophysically realistic network-level transport model of tau progression with exosome-mediated release and uptake processes

The spatiotemporal progression of tau in neurodegenerative diseases like Alzheimers follows the brains structural connectome, yet a gap exists between the macroscopic spread observed over years and the protein kinetics occurring over hours. Current models fail to reconcile this disparity or incorporate the cellular mechanisms driving transmission. Here, we advance the Network Transport Model (NTM) to bridge these scales by integrating active transport along microtubules, continuous toxic tau production, and exosome-mediated release and uptake. This framework constitutes one of the most biologically detailed models of tau spread on a whole brain to date, representing a significant innovation in how multiscale proteinopathies are simulated. Simulations on the mouse connectome demonstrate that this framework replicates empirical tau propagation patterns. Our results identify trans-neuronal release and uptake rates as the primary "bottleneck" on macroscopic spread, providing a biologically grounded explanation for the diseases slow progression. Furthermore, we find that high aggregation sequesters tau within regions, limiting global transmission, while increasing the abundance of toxic polymeric tau fibrils. Meanwhile, tau transport polarity bias (anterograde vs. retrograde) dictates spatial patterning. By linking molecular mechanics to system-wide pathology, this model provides an "in-silico" framework to evaluate how cellular-targeted interventions might alter the trajectory of tauopathic dementias. Author SummaryIn Alzheimers disease and related dementias, a protein called tau forms misfolded aggregates and gradually spreads through the brain along its neuronal wiring. A long-standing puzzle is linking the microscopic protein dynamics taking minutes to hours to the multiyear macroscopic spread of the disease. We developed a new mathematical model that is capable to explain macroscopic spreading behavior of tau as an emerging process from the underlying biophysical mechanics of tau pathology. Our simulations reproduce real patterns of tau spread, and our framework will pave the way for testing how treatments targeting specific cellular mechanisms might reshape disease progression.

neuroscience↗

bioRxiv2024

Multiplex genome editing eliminates the Warburg Effect without impacting growth rate in mammalian cells

The Warburg effect is ubiquitous in proliferative mammalian cells, including cancer cells, but poses challenges for biopharmaceutical production, as lactate accumulation inhibits cell growth and protein production. Previous efforts to eliminate lactate production via knockout have failed in mammalian bioprocessing since lactate dehydrogenase has proven essential. However, here we eliminated the Warburg effect in Chinese hamster ovary (CHO) and HEK293 cells by simultaneously knocking out lactate dehydrogenase and regulators involved in a negative feedback loop that typically inhibits pyruvate conversion to acetyl-CoA. In contrast to long-standing assumptions about the role of aerobic glycolysis, Warburg-null cells maintain wildtype growth rate while producing negligible lactate. Further characterization of Warburg-null CHO cells showed a compensatory increase in oxygen consumption, a near total reliance on oxidative metabolism, and higher cell densities in fed-batch cell culture. These cells remained amenable for production of diverse biotherapeutic proteins, reaching industrially relevant titers and maintaining product glycosylation. Thus, the ability to eliminate the Warburg effect is an important development for biotherapeutic production and provides a tool for investigating a near-universal metabolic phenomenon.

bioengineering↗

bioRxiv2022

Found in translation: Microproteins are a new class of potential host cell impurity in mAb drug products

Chinese hamster ovary (CHO) cells are used to produce almost 90% of therapeutic monoclonal antibodies (mAbs). The annotation of non-canonical translation events in these cellular factories remains incomplete, limiting not only our ability to study CHO cell biology but also detect host cell protein (HCP) contaminants in the final mAb drug product. We utilised ribosome footprint profiling (Ribo-seq) to identify novel open reading frames (ORFs) including N-terminal extensions and thousands of short ORFs (sORFs) predicted to encode microproteins. Mass spectrometry-based HCP analysis of four commercial mAb drug products using the extended protein sequence database revealed the presence of microprotein impurities for the first time. We also show that microprotein abundance varies with growth phase and can be affected by the cell culture environment. In addition, our work provides a vital resource to facilitate future studies of non-canonical translation as well as the regulation of protein synthesis in CHO cell lines.

genomics↗

bioRxiv2020

Tracing production instability in a clonally-derived CHO cell line using single cell transcriptomics

A variety of mechanisms including transcriptional silencing, gene copy loss and increased susceptibility to cellular stress have been associated with a sudden or gradual loss of monoclonal antibody (mAb) production in Chinese hamster ovary (CHO) cell lines. In this study, we utilised single cell RNA-seq (scRNA-seq) to study a clonally-derived CHO cell line that underwent production instability leading to a dramatic reduction of the levels of mAb produced. From the scRNA-seq data we identified sub clusters associated with variations in the mAb transgenes and observed that heavy chain gene expression was significantly lower than that of the light chain across the population. Using trajectory inference, the evolution of the cell line was reconstructed and was found to correlate with a reduction in heavy and light chain gene expression. Genes encoding for proteins involved in the response to oxidative stress and apoptosis were found to increase in expression as cells progressed along the trajectory. Future studies of CHO cell lines using this technology have the potential to dramatically enhance our understanding of the characteristics underpinning efficient manufacturing performance as well as product quality. HighlightsO_LIA clonally-derived CHO cell line in our laboratory had undergone production instability - in that the amount of intact monoclonal antibody had reduced dramatically to levels at which reliable quantitation was no longer possible. We were, however, able to detect mAb heavy and light chain protein, as well as dimerised light chain species in the cell culture media. C_LIO_LISingle cell RNA-seq was utilised to capture > 3,800 gene expression profiles from the cell line at 72hrs post seeding. C_LIO_LIAnalyses of the scRNA-seq data uncovered transcriptional heterogeneity and revealed the presence of multiple intra cell line clusters. The heavy chain transcript was detected at a significantly lower level in comparison to light chain transcripts. Light chain gene expression was not only more abundant, but also expressed more uniformly across the cell population. C_LIO_LIUsing unsupervised trajectory analysis, the emergence of heterogeneity in the cell population was traced from those cells most similar to the original isolated clone to those where transcription of the mAb heavy and light chain was undetectable. C_LIO_LISubsequent analysis of CHO cell gene expression patterns revealed a correlation between the progression of cells along the trajectory and the upregulation of genes involved in the cellular response to oxidative stress. C_LI

cell biology↗

bioRxiv2019

Sub physiological temperature induces pervasive alternative splicing in Chinese hamster ovary cells

RNA sequencing (RNASeq) has been widely used to associate alterations in Chinese hamster ovary (CHO) cell gene expression with bioprocess phenotypes, however alternative mRNA splicing, has thus far, received little attention. In this study, we utilised RNASeq for transcriptomic analysis of a monoclonal antibody producing CHOK1 cell line subjected to a temperature shift. More than 2,365 instances of differential splicing were observed 24hrs after the reduction of cell culture temperature. 1,163 of these alternative splicing events were identified in genes where no changes in abundance were detected by standard differential expression analysis. Ten examples of alternative splicing were selected for independent validation using qPCR in the monoclonal antibody producing CHOK1 line used for RNASeq and a further 2 CHOK1 cell lines. This analysis provided evidence that exon skipping and mutually exclusive splicing events occur in genes linked to the cellular response to changes in temperature and mitochondrial function. While further work is required to determine the impact of changes in mRNA sequence on cellular phenotype, this study demonstrates that alternative splicing analysis can be utilised to gain a deeper understanding of post-transcriptional regulation in CHO cells during biopharmaceutical production.

cell biology↗

bioRxiv2019

Expanding the Chinese hamster ovary cell long non-coding RNA transcriptome using RNASeq

Our ability to study Chinese hamster ovary (CHO) cell biology has been revolutionised over the last decade with the development of next generation sequencing and the publication of reference DNA sequences for CHO cells and the Chinese hamster. RNA sequencing has not only enabled the association of transcript expression with bioreactor conditions and desirable bioprocess phenotypes but played a key role in the characterisation of protein coding and small non-coding RNAs. The annotation of long non-coding RNAs, and therefore our understanding of their role in CHO cell biology, has been limited to date. In this manuscript, we use high resolution RNASeq data to more than double the number of annotated lncRNA transcripts for the CHOK1 genome. In addition, the utilisation of strand specific sequencing enabled the identification of more than 1,000 new lncRNAs located antisense to protein coding genes. The utility of monitoring lncRNA expression is demonstrated through an analysis of the transcriptomic response to a reduction of cell culture temperature and identification of simultaneous sense/antisense differential expression for the first time in CHO cells. To enable further studies of lncRNAs, the transcripts annotated in this study have been made available for the CHO cell biology community.

cell biology↗

bioRxiv2019

Cross-talk between m6A and m1A regulators, YTHDF2 and ALKBH3 fine-tunes mRNA expression

The recently re-discovered interest in N6-methyl adenosine (m6A) - one of more than a hundred modifications found on eukaryotic mRNA (known as epi-transcriptomic codes) - is currently one of the most topical areas in science. The m6A methylation impacts on all aspects of cellular RNA metabolism and several physiological processes. Although less abundant than the m6A epitranscriptomic mark, the m1A methylation has recently also attracted interest due to its dynamic nature in response to physiological changes. We investigated the role of the YTH domain-containing m6A reader protein family and the m1A eraser ALKBH3 on the expression of a transgene in mammalian cells. We present the first evidence that expression of a transgene is subjected to co-ordinated regulation by both m6A and m1A regulators. In addition, we provide genetic data implicating that the m6A reader YTHDF2 can read m1A site. Furthermore, we show that the m1A eraser ALKBH3 is a target of m1A methylation.

molecular biology↗