Biology subjects

Pyle, A.

Publications and source records attributed to Pyle, A..

5 recordsLinked to original sources

bioRxiv2026

Establishing the molecular basis for MDA5 mutation-linked autoimmunity

Melanoma differentiation-associated protein 5 (MDA5), a member of the RIG-I-like receptor family, is a cytoplasmic sensor essential for innate antiviral immunity. MDA5 distinguishes viral RNA from host RNA in part through its ATP hydrolysis activity, which promotes filament turnover on shorter endogenous dsRNAs. Here, we show that the gain-of-function T331I disease-linked mutation within the ATP binding pocket disrupts this balance, resulting in constitutive interferon signaling. Through a combination of cryo-electron microscopy (cryoEM), biochemical assays, and cellular analyses, we reveal the extensive network of interactions that precisely position ATP for catalysis in the wild-type MDA5 ATP binding pocket, and also demonstrate that the T331I mutation impairs ATPase activity, thereby stabilizing MDA5-dsRNA complexes and leading to aberrant immune activation. These findings elucidate how MDA5 ATPase activity regulates antiviral specificity and prevents autoimmunity by controlling filament stability and downstream signaling, offering a mechanistic molecular explanation for disease pathogenesis.

biochemistry↗

bioRxiv2024

Seed-competent alpha-synuclein pathology in metachromatic leukodystrophy: the expanding spectrum of alpha-synucleinopathy in sphingolipidoses

Metachromatic leukodystrophy (MLD) is a rare - typically paediatric - sphingolipid storage disorder resulting from bi-allelic pathogenic variants in the ARSA gene, encoding the lysosomal arylsulphatase A (ASA). Heterozygous variants in ARSA are associated with risk of Lewy body diseases (LBD), a group of age-associated neurodegenerative disorders characterised by the accumulation of the protein -synuclein; however, no study has yet determined whether -synuclein with putative pathological features is observed in MLD brain tissue. We examined post-mortem brain tissue from MLD cases (N=5, age 2-33) compared to matched control cases using histological approaches and -synuclein seeding amplification assay (SAA). Juvenile-onset MLD cases exhibited granular -synuclein deposits in neurons of regions prone to neuronal pathology in MLD, and seed-competent conformers that generated atypical short, twisted fibrils on SAA. In contrast, infantile-onset MLD cases gave only variably positive reactions on SAA. In summary, this study suggests MLD cases manifest -synuclein pathology reminiscent of that observed in LBD, even in juvenile populations, further expanding the spectrum of sphingolipid storage disorders associated with the aggregation of -synuclein. These findings have important implications for understanding the disease process of both LBD and MLD, potentially highlighting novel pathways for therapeutic interventions in both conditions.

neuroscience↗

bioRxiv2023

Nanobiopsy investigation of the subcellular mtDNA heteroplasmy in human tissues

Mitochondrial function is critical to continued cellular vitality and is an important contributor to a growing number of human diseases. Mitochondrial dysfunction is typically heterogeneous, mediated through the clonal expansion of mitochondrial DNA (mtDNA) variants in a subset of cells in a given tissue. To date, our understanding of the dynamics of clonal expansion of mtDNA variants has been technically limited to the single cell-level. Here, we report the use of nanobiopsy for subcellular sampling from human tissue, combined with next-generation sequencing to assess subcellular mtDNA mutation load in human tissue from mitochondrial disease patients. The ability to map mitochondrial mutation loads within individual cells of diseased tissue samples will further our understanding of mitochondrial genetic diseases.

genomics↗

bioRxiv2021

A subcellular cookie cutter for spatial genomics in human tissue

Intracellular heterogeneity contributes significantly to cellular physiology and, in a number of debilitating diseases, cellular pathophysiology. This is greatly influenced by distinct organelle populations and to understand the aetiology of disease it is important to have tools able to isolate and differentially analyse organelles from precise location within tissues. Here we report the development of a subcellular biopsy technology that facilitates the isolation of organelles, such as mitochondria, from human tissue. We compared the subcellular biopsy technology to laser capture microdissection (LCM) that is the state of art technique for the isolation of cells from their surrounding tissues. We demonstrate an operational limit of (>20m) for LCM and then, for the first time in human tissue, show that subcellular biopsy can be used to isolate mitochondria beyond this limit.

genomics↗

bioRxiv2018

A genome-wide association study of mitochondrial DNA copy number in two population-based cohorts

Mitochondrial DNA copy number (mtDNA CN) exhibits interindividual and intercellular variation, but few genome-wide association studies (GWAS) of directly assayed mtDNA CN exist.\n\nWe undertook a GWAS of qPCR-assayed mtDNA CN in the Avon Longitudinal Study of Parents and Children (ALSPAC), and the UK Blood Service (UKBS) cohort. After validating and harmonising data, 5461 ALSPAC mothers (16-43 years at mtDNA CN assay), and 1338 UKBS females (17-69 years) were included in a meta-analysis. Sensitivity analyses restricted to females with white cell-extracted DNA, and adjusted for estimated or assayed cell proportions. Associations were also explored in ALSPAC children, and UKBS males.\n\nA neutrophil-associated locus approached genome-wide significance (rs709591 [MED24], {beta}[SE] -0.084 [0.016], p=1.54e-07) in the main meta-analysis of adult females. This association was concordant in magnitude and direction in UKBS males and ALSPAC neonates. SNPs in and around ABHD8 were associated with mtDNA CN in ALSPAC neonates (rs10424198, {beta}[SE] 0.262 [0.034], p=1.40e-14), but not other study groups. In a meta-analysis of unrelated individuals (N=11253), we replicated a published association in TFAM {beta}[SE] 0.046 [0.017], p=0.006), with an effect size much smaller than that observed in the replication analysis of a previous in silico GWAS.\n\nIn a hypothesis-generating GWAS, we confirm an association between TFAM and mtDNA CN, and present putative loci requiring replication in much larger samples. We discuss the limitations of our work, in terms of measurement error and cellular heterogeneity, and highlight the need for larger studies to better understand nuclear genomic control of mtDNA copy number.

genomics↗