Biology subjects

Hudson, D.

Publications and source records attributed to Hudson, D..

6 recordsLinked to original sources

bioRxiv2026

Reverse engineering the fatally cross-reactive A3A TCR to decouple potency and specificity

T cell receptor (TCR) affinity enhancement can introduce off-target cross-reactivity with life-threatening consequences, as illustrated by the MAGE-A3-specific A3A TCR, which caused fatal cardiotoxicity through recognition of a Titin-derived peptide. Here, we reconstructed the cross-reactivity landscape by reverse-engineering A3A toward its wild-type precursor, generating intermediate variants in which engineered CDR2 residues are systematically reverted to the wild-type sequence. Reverting just two engineered residues yields a receptor, v9, that retains MAGE-A3 cytotoxicity comparable to A3A while eliminating Titin and other acquired cross-reactivities. Structurally, these substitutions reduce CDR2-MHC contacts and disrupt an intra-TCR CDR2-CDR3{beta} interaction, propagating conformational changes across CDR3 loops that reshape peptide engagement without altering docking geometry. These results demonstrate that mutations outside the peptide-contacting CDR3 loops can allosterically reconfigure antigen specificity and establish simple stepwise reverse engineering to wild-type as a strategy for correcting TCR cross-reactivity.

immunology↗

bioRxiv2023

The role of family and environment in determining the skin microbiome of captive aquatic frogs, Xenopus laevis

BackgroundThe amphibian skin microbiome has drawn interest due to the ecological threat posed by chytridiomycosis, which drives changes in symbiotic microbial communities and may be inhibited by certain bacterial taxa. However, skin microbes also play a role in amphibian tissue regeneration. Xenopus spp. are well-established model organisms used to study development, regeneration, genetics and disease. Husbandry protocols, including use of antibiotics and other sterilising agents, may affect experimental outcomes by altering microbiomes. It is therefore essential to improve our understanding of Xenopus microbiome characteristics and inheritance. We undertook bacterial 16S rRNA based sampling of a captive, closed Xenopus laevis colony. A total of 16 female frogs, their eggs, and tadpoles were sampled, covering multiple aquarium systems and tanks, along with testes from males used for in vitro fertilisation and a range of environmental samples. ResultsTank environments supported the most complex microbial communities. Mother frogs harboured the most diverse microbial communities of the frog life stages, with tadpole skin microbiomes being relatively simple. Frog samples were dominated by Proteobacteria and Bacteroidota. Rhizobium and Chryseobacterium were dominant in tadpoles, whereas mothers supported high proportions of Vogesella and Acinetobacter as well as Chryseobacterium. While the mothers habitats contained low levels of these taxa, the tadpoles environmental microbes were very similar to those on tadpole skin. A total of 34 genera were found to be differentially abundant between the mothers and tadpoles. Analysis of Bray-Curtis distances indicated that mother and tadpole microbiomes varied according to the mothers aquarium system, the tanks within them, and the individual mother. Source tracking analyses showed that egg jelly and tadpoles received a mean of approximately two thirds of their microbiomes via vertical transmission, although a sizeable proportion came from unknown sources at all life stages. ConclusionsThe skin of mother frogs appears to select for certain taxa that are otherwise present at low abundances in the environment. While tadpoles inherit a proportion of their microbiomes from their mothers via the egg, they support a distinct and less diverse microbial community than adult frogs. The microbiome varies between individual mothers, and is also affected by the aquarium system and individual tank within that the mother occupies.

microbiology↗

bioRxiv2023

A comparison of clustering models for inference of T cell receptor antigen specificity

The vast potential sequence diversity of TCRs and their ligands has presented an historic barrier to computational prediction of TCR epitope specificity, a holy grail of quantitative immunology. One common approach is to cluster sequences together, on the assumption that similar receptors bind similar epitopes. Here, we provide an independent evaluation of widely used clustering algorithms for TCR specificity inference, observing some variability in predictive performance between models, and marked differences in scalability. Despite these differences, we find that different algorithms produce clusters with high degrees of similarity for receptors recognising the same epitope. Our analysis highlights an unmet need for improvement of complex models over a simple Hamming distance comparator, and strengthens the case for use of clustering models in TCR specificity inference.

immunology↗

bioRxiv2021

CRISPR-Cas9 gene editing and rapid detection of gene-edited mutants using high-resolution melting in the apple scab fungus, Venturia inaequalis

BackgroundScab, or black spot, caused by the filamentous fungal pathogen Venturia inaequalis, is the most economically important disease of apple (Malus x domestica) worldwide. To develop durable control strategies against this disease, a better understanding of the genetic mechanisms underlying the growth, reproduction, virulence and pathogenicity of V. inaequalis is required. A major bottleneck for the genetic characterization of V. inaequalis is the inability to easily delete or disrupt genes of interest using homologous recombination. Indeed, no gene deletions or disruptions in V. inaequalis have yet been published. Recently, CRISPR-Cas9 has emerged as an efficient tool for gene editing in filamentous fungi. With this in mind, we set out to establish CRISPR-Cas9 as a gene editing tool in V. inaequalis. ResultsWe showed that CRISPR-Cas9 can be used for gene inactivation in the apple scab fungus. As a proof of concept, we targeted the melanin biosynthesis pathway gene trihydroxynaphthalene reductase (THN), which has previously been shown to result in a light-brown colony phenotype when transcriptionally silenced using RNA interference. Using one of two CRISPR-Cas9 single guide RNAs (sgRNAs) targeted to the THN gene, delivered by a single autonomously replicating Golden Gate-compatible plasmid, we were able to identify six of 36 stable transformants with a light-brown phenotype, indicating an ~16.7% gene inactivation efficiency. Notably, of these six THN mutants, five had an independent mutation. As part of our pipeline, we also report a high-resolution melting (HRM) curve protocol for the rapid detection of CRISPR-Cas9 gene-edited mutants of V. inaequalis. This protocol identified a single base pair deletion mutation in a sample containing only 5% mutant genomic DNA, indicating high sensitivity for mutant screening. ConclusionsIn establishing CRISPR-Cas9 as a tool for gene editing in V. inaequalis, we have provided a strong starting point for studies aiming to decipher the function of genes associated with the growth, reproduction, virulence and pathogenicity of this fungus. The associated HRM curve protocol will enable CRISPR-Cas9 transformants to be screened for gene inactivation in a high-throughput and low-cost manner, which will be particularly powerful in cases where the CRISPR-Cas9-mediated gene inactivation efficiency is low.

microbiology↗

bioRxiv2020

Quantifying the wave resistance of a swimmer

AbstractQuantifying the wave resistance of a swimmer as a function of depth assists in identifying the optimum depth for the glide phases of competition. Previous experiments have inferred how immersed depth influences the drag acting on a swimmer [1], but have not directly quantified the magnitude of wave resistance. This research experimentally validates the use of thin-ship theory for quantifying the wave resistance of a realistic swimmer geometry. The drag and wave pattern of a female swimmer mannequin were experimentally measured over a range of depths from 0.05m to 1.00m at a speed of 2.50 m/s. Numerical simulations agree with experiment to confirm that there were negligible reductions in wave resistance below a depth of 0.40m. Larger swimming pool dimensions are shown to be significant at reducing wave resistance at speeds above 2.0 m/s and depths below 0.40m. Truncating the swimmer’s body at the upper thigh increases the wave resistance at speeds below 2.0m/s but is not significant at higher speeds, indicating that the upper body is the main contributor to the wave system. Numerical experiments indicate that rotating the shoulders towards the surface is more influential than the feet, demonstrating the impact of the upper body on wave resistance.Competing Interest StatementThe authors have declared no competing interest.View Full Text

bioengineering↗

bioRxiv2020

Tendon and Motor Phenotypes in the Crtap-/- Mouse Model of Recessive Osteogenesis Imperfecta

Osteogenesis imperfecta (OI) is characterized by short stature, skeletal deformities, low bone mass with bone fragility, and motor deficits. A subset of OI patients also present with joint hypermobility; however, the role of tendon/ligament dysfunction in OI pathogenesis is largely unknown. Using the Crtap-/- mouse model of severe, recessive OI, we found that mutant Achilles tendons and patellar ligaments were thinner with increased collagen cross-links and reduced collagen fibril size at 1- and 4-months compared to wildtype. Patellar ligaments from Crtap-/- mice also had fewer progenitors with a concomitant increase in immature cells. RNA-seq analysis of Achilles tendons and patellar ligaments from 1-month Crtap-/- mice revealed dysregulation in matrix gene expression concomitant with predicted alterations in TGF-{beta}, inflammatory, and metabolic signaling. Finally, a series of behavioral tests revealed severe motor impairments and reduced grip strength in 4-month Crtap-/- mice - a phenotype that correlates with the tendon/ligament pathology.

genetics↗