Biology subjects

Burns, C.

Publications and source records attributed to Burns, C..

14 recordsLinked to original sources

bioRxiv2026

Mucosal-associated invariant T cells support IL-15-dependent Treg response in skin injury and promote resolution of skin inflammation

Mucosal-associated invariant T (MAIT) cells are enriched at barrier sites, but their role in autoimmune skin inflammation remains unknown. Using cutaneous lupus as a model, we identify MAIT cells as protective regulators of skin inflammation and as critical upstream modulators of regulatory T cells (Treg). Topical MAIT cell activation with 5-OP-RU induced durable resolution of spontaneous skin lesions in MRL/lpr mice and suppressed TLR7-driven skin inflammation. MAIT cell activation selectively expanded and activated Treg populations in both healthy and lupus-like skin, while suppressing effector T cell cytokine production and cytotoxic programs. This MAIT-Treg axis was also activated in UV light-driven barrier injury in healthy murine and human skin, where MAIT cells were required for UV-elicited Treg expansion and function. In lupus-like skin, local MAIT cell activation restored the defective UVB-induced Treg response and limited CD8+ T cell expansion. Mechanistically, CCR2+ monocyte-derived antigen-presenting cells and IL-15 signaling were required for MAIT cell-driven Treg accumulation and therapeutic benefits of MAIT cells in inflamed skin. These studies identify a MAIT-IL-15-Treg axis that links barrier injury sensing to immune regulation, which is disrupted in cutaneous lupus, and nominate therapeutic MAIT cell activation as an unappreciated strategy for restoring immune homeostasis in inflamed skin.

immunology↗

bioRxiv2025

A comprehensive genetic toolkit for Zymomonas mobilis; allowing rapid, high-resolution, combinatorial engineering of metabolic pathways.

The limited availability of reliable genetic tools has constrained the precise engineering of Zymomonas mobilis, a promising bacterium for the sustainable bioproduction of fuels and commodity chemicals. Here, we present a comprehensive, modular genetic toolkit that enables high-resolution, combinatorial control of gene expression in Z. mobilis. We constructed and characterised a synthetic promoter library spanning the broadest dynamic range reported for this organism, alongside a suite of strong transcriptional terminators that effectively insulate genetic cassettes and prevent transcriptional readthrough. To achieve predictable translation across diverse genetic contexts, we further implemented bicistronic design (BCD) elements, allowing context-independent control of expression. Promoter activities were shown to be stable under both aerobic and anaerobic conditions, highlighting the robustness of the regulatory parts across physiologically relevant environments. The toolkit was integrated into the Start-Stop Assembly system, facilitating high-throughput construction of multi-gene pathways. We demonstrate its utility through the rapid assembly and testing of the possible expression space of genes encoding a 2,3-butanediol biosynthetic pathway. Collectively, this genetic toolkit now allows the high-resolution engineering of multi-enzyme pathways in Z. mobilis for industrial biotechnology applications.

synthetic biology↗

bioRxiv2024

Semaphorin 4A maintains functional diversity of the hematopoietic stem cell pool

Somatic stem cell pools are comprised of diverse, highly specialized subsets whose individual contribution is critical for the overall regenerative function. In the bone marrow, myeloid-biased HSC (myHSC) are indispensable for replenishment of myeloid cells and platelets during inflammatory response but at the same time, become irreversibly damaged during inflammation and aging. Here, we identify an extrinsic factor, Semaphorin 4A (Sema4A), which non cell-autonomously confers myHSC resilience to inflammatory stress. We show that the absence of Sema4A, myHSC inflammatory hyper-responsiveness in young mice drives excessive myHSC expansion, myeloid bias and profound loss of regenerative function with age. Mechanistically, Sema4A is mainly produced by neutrophils, signals via a cell surface receptor Plexin D1 and safeguards myHSC epigenetic state. Our study shows that by selectively protecting a distinct stem cell subset, an extrinsic factor preserves functional diversity of somatic stem cell pool throughout organismal lifespan.

cell biology↗

bioRxiv2024

Heterogeneity and plasticity of the naive CD4+ T cell compartment

While naive CD4+ T cells have historically been considered a homogenous population, recent studies have provided evidence that functional heterogeneity exists within this population. Using single cell RNA sequencing (scRNAseq), we identify five transcriptionally distinct naive CD4+ T cell subsets that emerge within the single positive stage in the thymus: a quiescence cluster (TQ), a memory-like cluster (TMEM), a TCR reactive cluster (TTCR), an IFN responsive cluster (TIFN), and an undifferentiated cluster (TUND). Elevated expression of transcription factors KLF2, Mx1, and Nur77 within the TQ, TIFN, and TMEM clusters, respectively, allowed enrichment of these subsets for further analyses. Functional studies using sorted cells revealed that naive T cell subsets have distinctive functional biases upon stimulation. Furthermore, treatment of mice with inflammatory stimuli imparted a state of reduced responsiveness on naive T cells, evidenced by a reduction in cytokine production ex vivo. In human lupus patients, naive CD4+ T cell cluster frequencies were distorted, with the TIFN cluster expanding proportionately with disease score. Our data show that naive T cells are influenced by host environment, with functional consequences manifesting upon activation. These findings highlight a need to explore how naive T cells can become distorted in cancer, autoimmunity, and infectious diseases. SummaryThis study describes the transcriptional heterogeneity of murine and human naive CD4+ T cells as comprising of multiple discrete clusters that impact CD4+ T cell fate and trajectories. Naive CD4+ T cells experiencing inflammatory environments exhibit an altered transcriptional state that influences their functional trajectory.

immunology↗

bioRxiv2024

Loss of VPS41 triggers rapid insulin degradation and dysregulated autophagy in pancreatic beta-cells

Vacuolar protein sorting-associated protein 41 (VPS41) has been established as a requirement for normal insulin secretory function in pancreatic beta-cells. Genetic deletion of VPS41 in mouse pancreatic beta-cells results in diabetes, though the mechanisms are not understood. Presently, we show that VPS41 deletion results in rapid mature insulin degradation and downregulation of beta-cell identity. This phenotype is observed in vivo, with VPS41KO mice displaying progressive loss of insulin content and beta-cell function with age. In acute VPS41 depletion in vitro, the loss of insulin is associated with increased degradative pathway activity, increased Adapter Protein 3 complex colocalisation with lysosomes, increased nuclear localisation of transcription factor E3, and downregulation of PDX1 and INS mRNA expression. Inhibition of lysosomal degradation rescues the rapidly depleted insulin content. These data evidence a VPS41-dependent mechanism for both insulin content degradation and loss of beta-cell identity in beta-cells.

cell biology↗

bioRxiv2022

Thymic mesenchymal niche cells drive T cell immune regeneration

Thymic atrophy and the progressive immune decline that accompanies it is a major health problem, chronically with age and acutely with immune injury. No solution has been defined. Here we demonstrate that one of the three mesenchymal cell subsets identified by single-cell analysis of human and mouse thymic stroma is a critical niche component for T lymphopoiesis. The Postn+ subset is located perivascularly in the cortical-medullary junction, medulla and subcapsular regions. Cell depletion demonstrated that it recruits T competent cells to the thymus and initiates T lymphopoiesis in vivo. This subset distinctively expresses the chemokine Ccl19 necessary for niche functions. It markedly declines with age and in the acute setting of hematopoietic stem cell transplant conditioning. When isolated and adoptively transferred, these cells durably engrafted the atrophic thymus, recruited early T progenitors, increased T cell neogenesis, expanded TCR complexity and enhanced T cell response to vaccination. These data define a thymus lymphopoietic niche cell type that may be manipulated therapeutically to regenerate T lymphopoiesis.

immunology↗

bioRxiv2022

Towards optimised extracellular vesicle proteomics from cerebrospinal fluid

The proteomic profile of extracellular vesicles (EVs) from cerebrospinal fluid (CSF) can reveal novel biomarkers for diseases of the brain. Here, we validate an ultrafiltration combined with size-exclusion chromatography (UF-SEC) method for isolation of EVs from canine CSF and probe the effect of starting volume on the EV proteomics profile. First, we performed a literature review of CSF EV articles to define the current state of art, discovering a need for basic characterisation of CSF EVs. Secondly, we isolated EVs from CSF by UF-SEC and characterised the SEC fractions by protein amount, particle count, transmission electron microscopy, and immunoblotting. Data are presented as mean {+/-} standard deviation. Using proteomics, SEC fractions 3-5 were compared and enrichment of EV markers in fraction 3 was detected, whereas fractions 4-5 contained more apolipoproteins. Lastly, we compared starting volumes of pooled CSF (6ml, 3ml, 1ml, and 0.5ml) to evaluate the effect on the proteomic profile. Even with a 0.5ml starting volume, 743{+/-}77 or 345{+/-}88 proteins were identified depending on whether matches between runs was active in MaxQuant. The results confirm that UF-SEC effectively isolates CSF EVs and that EV proteomic analysis can be performed from 0.5ml of canine CSF.

cell biology↗

bioRxiv2022

Myeloid-biased HSC require Semaphorin 4A from the bone marrow niche for self-renewal under stress and life-long persistence

Tissue stem cells are hierarchically organized. Those that are most primitive serve as key drivers of regenerative response but the signals that selectively preserve their functional integrity are largely unknown. Here, we identify a secreted factor, Semaphorin 4A (Sema4A), as a specific regulator of myeloid-biased hematopoietic stem cells (myHSC), which are positioned at the top of the HSC hierarchy. Lack of Sema4A leads to exaggerated myHSC (but not downstream "balanced" HSC) proliferation after acute inflammatory stress, indicating that Sema4A enforces myHSC quiescence. Strikingly, aged Sema4A knock-out myHSC expand but almost completely lose reconstitution capacity. The effect of Sema4A is non cell-autonomous, since upon transplantation into Sema4A-deficient environment, wild-type myHSC excessively proliferate but fail to engraft long-term. Sema4A constrains inflammatory signaling in myHSC and acts via a surface receptor Plexin-D1. Our data support a model whereby the most primitive tissue stem cells critically rely on a dedicated signal from the niche for self-renewal and life-long persistence.

cell biology↗

bioRxiv2021

Deletion of Vhl in Dmp1-expressing cells causes microenvironmental impairment of B cell lymphopoiesis

The contributions of skeletal cells to the processes of B cell development in the bone marrow (BM) have not been completely described. The von-Hippel Lindau protein (VHL) plays a key role in cellular responses to hypoxia. Previous work showed that Dmp1-Cre;Vhl conditional knockout mice (VhlcKO), which delete Vhl in late osteoblasts and osteocytes, display dysregulated bone growth and reduction in B cells. Here, we investigated the mechanisms underlying the B cell defects using flow cytometry and high-resolution imaging. In the VhlcKO BM, B cell progenitors were increased in frequency and number, whereas Hardy Fractions B-F were decreased. VhlcKO Fractions B-C cells showed increased apoptosis and quiescence. Reciprocal BM chimeras confirmed a B cell-extrinsic source of the VhlcKO B cell defects. In support of this, VhlcKO BM serum contained reduced CXCL12 and elevated EPO levels. Staining of VhlcKO B cells with an intracellular hypoxic marker indicated the natural existence of distinct B cell microenvironments that differ in local oxygen tensions. Additionally, intravital and ex vivo imaging revealed VhlcKO BM blood vessels with increased diameter, frequency, volume, and a diminished blood-BM barrier. Our studies identify novel mechanisms linking altered bone homeostasis with drastic BM microenvironmental changes that dysregulate B cell development.

immunology↗

bioRxiv2020

Traumatic stress history interacts with chronic peripheral inflammation to alter mitochondrial function of synaptosomes in a sex-specific manner

BackgroundRepeated exposures to chronic stress can lead to long lasting negative behavioral and metabolic outcomes. Here, we aim to determine the impact of chronic stress and chronic low-level inflammation on behavior and synaptosomal metabolism. MethodsMale (n = 31) and female (n = 32) C57Bl/6 mice underwent chronic repeated predation stress or daily handling for two rounds of 15 consecutive days of exposure during the adolescent and early adult timeframes. Subsequently, mice were exposed to repeated lipopolysaccharide (LPS; 7.5 x 105 EU/kg) or saline injections every third day for eight weeks. Exploratory and social behaviors were assessed in the open field and social interaction tests prior to examination of learning and memory with the Barnes Maze. Mitochondrial function and morphology were assessed in synaptosomes post-mortem. In addition, expression of TNF-, IL-1{beta}, and ROMO1 were examined in the hippocampus and prefrontal cortex. Circulating pro- and anti-inflammatory cytokines in the periphery were assessed following the first and last LPS injection as well as at the time of tissue collection. Circulating ROMO1 was assessed in terminal samples. ResultsExposure to repeated predatory stress increased time spent in the corners of the open field, suggestive of anxiety-like behavior, in both males and females. There were no significant group differences in the social interaction test and minimal effects were evident in the Barnes maze. A history of chronic stress interacted with chronic LPS in male mice to lead to a deficit in synaptosomal respiration. Female mice were more sensitive to both chronic stress and chronic LPS such that either a history of chronic stress or a history of chronic LPS was sufficient to disrupt synaptosomal respiration in females. Both stress and chronic LPS were sufficient to increase inflammation and reactive oxygen in males in the periphery and centrally. Females had increased markers of peripheral inflammation following acute LPS but no evidence of peripheral or central increases in inflammatory factors or reactive oxygen following chronic exposures. ConclusionCollectively, these data suggest that while metrics of inflammation and reactive oxygen are disrupted in males following chronic stress and chronic LPS, only the combined condition is sufficient to alter synaptosomal respiration. Conversely, although evidence of chronic inflammation or chronic elevation in reactive oxygen is absent, females demonstrate profound shifts in synaptosomal mitochondrial function with either a history of chronic stress or a history of chronic inflammation. These data highlight that differential mechanisms are likely in play between the sexes and suggest that female sensitivity to neurogenerative conditions may be precipitated by influence of life experiences on mitochondrial function in the synapses.

neuroscience↗

bioRxiv2019

VPS41 recessive mutation causes ataxia and dystonia with retinal dystrophy and mental retardation by inhibiting HOPS function and mTORC1 signaling

The vacuolar protein sorting protein 41 (VPS41) is a neuroprotective protein in models of Parkinsons disease (PD). As part of the HOPS (Homotypic fusion and Protein Sorting) complex, VPS41 regulates fusion of lysosomes with late endosomes and autophagosomes. Independent of HOPS, VPS41 regulates transport of newly synthesized lysosomal membrane proteins and secretory proteins. Here we report two brothers with compound heterozygous mutations in VPS41 (VPS41R662* and VPS41S285P), born to healthy and non-consanguineous parents. Both patients displayed transient retinal dystrophy, ataxia and dystonia, with brain MRI findings of cerebellar atrophy and a thin saber-shape corpus callosum. Patient-derived fibroblasts contained enzymatically active lysosomes that were poorly reached by endocytic cargo and failed to attract the mTORC1 complex. Consequently, transcription factor TFE3, a driver of autophagy and lysosomal genes, showed continuous nuclear localization which resulted in elevated LC3-II levels and an impaired response to nutrient starvation. CRISPR/CAS VPS41 HeLa knockout cells showed a similar phenotype that could be rescued by wildtype VPS41 but not by VPS41S285P or VPS41R662*. mTORC1 inhibition was also seen after knockout of HOPS subunits VPS11 or VPS18. Regulated neuropeptide secretion in PC12 VPS41 knockout cells was rescued by VPS41S285P expression, indicating that this HOPS-independent function was preserved. Co-expression of the VPS41S285P and VPS41R662* variants in a C. elegans model of PD abolished the protective effect of VPS41 against -synuclein-induced neurodegeneration. We conclude that both disease-associated VPS41 variants specifically abrogate HOPS function, which leads to a delay in endocytic cargo delivery to lysosomes, mTORC1 inhibition and irresponsiveness to autophagic clues. Our studies signify a link between HOPS function and mTORC1 signaling and imply that HOPS function is required for the neuroprotective effect of VPS41 in PD.

cell biology↗

bioRxiv2019

Methods for combining multiple correlated biomarkers with application to the study of low-grade inflammation and muscle mass in senior horses

The simplest analysis of biomarker data is based on a series of single biomarker hypothesis tests, followed by correction for multiple testing. However, it is intuitively plausible that a joint analysis of multiple biomarkers will have higher statistical power and promise improved discrimination over tests based on single markers. In this article, we study analytical properties of the approach for joint analysis of correlated summary statistics based on the test for quadratic forms (TQ). Based on the derivation of the TQ-distribution, we proposed a scale-location approximation of the TQ statistic, which we call approximate TQ. We show that the approximate TQ has very similar power to the traditional TQ test under varying correlation structures among biomarkers. Our application of both the TQ and the approximate TQ test to data on biomarkers for inflamm-aging - an age-related low-grade chronic inflammation - reveals an association between the percentage of IFN{gamma} positive lymphocytes and overall muscle condition in senior horses.

genetics↗

bioRxiv2018

Bud13 Promotes a Type I Interferon Response By Countering Intron Retention in Irf7

Intron retention (IR) has emerged as an important mechanism of gene expression control. Despite this, the factors that control IR events remain poorly understood. We observed consistent IR in one intron of the Irf7 gene and identified Bud13 as an RNA-binding protein that acts at this intron to increase the amount of successful splicing. Deficiency in Bud13 led to increased IR, decreased mature Irf7 transcript and protein levels, and consequently to a dampened type I interferon response. This impairment of Irf7 production in Bud13-deficient cells compromised their ability to withstand VSV infection. Global analysis of Bud13 knockdown and BUD13 cross-linking to RNA revealed a subset of introns that share many characteristics with the one found in Irf7 and are spliced in a Bud13-dependent manner. Deficiency of Bud13 led to decreased mature transcript from genes containing such introns. Thus, by acting as an antagonist to IR, Bud13 facilitates the expression of genes at which IR occurs.

molecular biology↗

bioRxiv2017

HID-1 controls cargo sorting and dense core formation by influencing trans- Golgi network acidification in neuroendocrine cells

Large dense core vesicles (LDCVs) mediate the regulated release of neuropeptides and peptide hormones. They form at the trans-Golgi network (TGN) where their soluble content aggregates to form a dense core, but the mechanisms controlling biogenesis are still not completely understood. Recent studies have implicated the peripheral membrane protein HID-1 in neuropeptide sorting and insulin secretion. Using CRISPR/Cas9, we generated HID-1 KO rat neuroendocrine cells, and show that the absence of HID-1 results in specific defects in peptide hormone and monoamine storage and regulated secretion. Loss of HID-1 causes a reduction in the number of LDCVs and affects their morphology and biochemical properties due to impaired cargo sorting and dense core formation. HID-1 KO cells also exhibit defects in TGN acidification together with mislocalization of the Golgi-enriched vacuolar H+-ATPase subunit isoform a2. We propose that HID-1 influences early steps in LDCV formation by controlling dense core formation at the TGN.

cell biology↗