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Salio, M.

Publications and source records attributed to Salio, M..

9 recordsLinked to original sources

Neonatal AAV-SIL1 gene therapy prevents Marinesco-Sjögren syndrome in mice

Marinesco-Sjogren syndrome is a rare early-onset multisystem disorder characterized primarily by cerebellar ataxia and myopathy and caused by loss-of-function mutations in SIL1. No disease-modifying therapy is available. We investigated whether adeno-associated virus (AAV)-mediated gene therapy could prevent neurological and muscular disease in the woozy mouse model of SIL1 deficiency. Neonatal mice received intracerebroventricular injections of AAV-PHP.eB vectors expressing SIL1 under the control of either a ubiquitous or a Purkinje cell-specific promoter. Vehicle-treated woozy mice developed progressive motor impairment accompanied by extensive Purkinje cell degeneration, thinning of the cerebellar molecular layer, marked astrogliosis and activation of endoplasmic reticulum stress pathways. In contrast, AAV-SIL1 treatment prevented the onset of ataxia and substantially preserved cerebellar architecture, including Purkinje cells and molecular-layer thickness, while reducing astrogliosis and endoplasmic reticulum stress. Selective restoration of SIL1 expression in Purkinje cells was sufficient to rescue motor performance, supporting a major cell-autonomous contribution of Purkinje cell dysfunction to cerebellar disease. Therapeutic benefit was maintained throughout the 26-week observation period, the longest time point examined, with treated mice remaining behaviorally indistinguishable from heterozygous controls. AAV-SIL1 treatment also improved muscle function and markedly attenuated skeletal muscle pathology, with transgenic SIL1 expression in muscle reaching levels comparable to those in heterozygous controls. These findings provide proof-of-principle that AAV-mediated SIL1 gene therapy can prevent the neurological and muscular manifestations of Marinesco-Sjogren syndrome and identify Purkinje cells as a critical cellular target for preventing cerebellar dysfunction.

neuroscience↗

Gut archaea drive human MAIT cell responses

Archaea, the third domain of life, are a gut microbiome constituent, but their role in shaping immunity is poorly understood. Mucosal-associated invariant T (MAIT) cells sense microbial metabolites through their T cell receptor and contribute to antimicrobial defence and barrier homeostasis. Key sources of the major MAIT cell antigen 5-OP-RU within the human gut microbiome are unknown. Analysing microbiomes from UK, Zambian, Kenyan and Ugandan donors, we found strong correlations between archaeal riboflavin biosynthesis and circulating MAIT cell frequency and activation. Stool 5-OP-RU levels correlated with archaeal abundance. Cultures of methanogenic archaea, including the human commensal Methanobrevibacter smithii, produced 5-OP-RU and strongly activated MAIT cells without inflammation. We demonstrate a critical role for archaea in shaping human immunity, with implications for microbiome-mediated regulation of health.

immunology↗

ImmTACs overcome cytotoxic T cell suppression

Immune mobilizing monoclonal TCR against cancer (ImmTAC) are cancer therapeutics that activate T cells through recognition of a tumor-associated antigenic MHC/peptide complex. A first-in-class ImmTAC, Tebentafusp, is approved for the treatment of metastatic uveal melanoma. While clinical efficacy is thus established, the cellular mechanisms underpinning ImmTAC action are not fully resolved. Using a recently established experimental strategy to generate suppressed human primary cytotoxic T lymphocytes (CTL), we have investigated an ImmTAC that recognizes a peptide derived from the tumor associated antigen NY-ESO-1 in comparison to direct engagement of a TCR recognizing the same MHC/peptide complex. In response to endogenous antigen presentation, ImmTACs could elicit tumor cell cytolysis by suppressed CTL, but not IFN{gamma} secretion, in a manner dependent on the engager affinity for CD3{varepsilon}. ImmTACs enhanced the efficient execution of subcellular CTL polarization steps required for effective cytolysis and could trigger calcium signaling. These data establish that ImmTACs activate CTL similarly to direct engagement of a TCR by MHC/peptide and are likely to retain this capability under suppressive conditions such as in the tumor microenvironment.

immunology↗

Ligand-responsive groove remodelling in human and macaque CD1d reveals a conserved MHC-like gating mechanism

CD1d presents lipid antigens to invariant natural killer T (iNKT) cells. We determined a high-resolution crystal structure of human CD1d bound to -galactosylceramide (-GalCer) at 1.76 [A], enabling detailed investigation of ligand-sensitive conformational flexibility at Phe84, a conserved aromatic residue that caps the F' groove. Electron density at Phe84 revealed multiple side-chain conformations, suggestive of ligand-induced plasticity. Molecular dynamics simulations indicated that the canonical rotamer is energetically favoured in the absence of a stabilising groove-occupying ligand. To assess conservation of this putative gating mechanism, we solved the first CD1d structure from a non-human primate, rhesus macaque CD1d--GalCer, at 1.83 [A] resolution. In contrast to the human complex, Phe84 in macaque CD1d adopted a fixed conformation. As this aromatic residue is conserved across CD1 isoforms and CD1d-expressing species, and mirrors gating residues in MHC class I that regulate peptide accommodation, our findings support a shared evolutionary strategy for managing antigen diversity. These data provide critical insight into the mechanisms of antigen presentation by CD1 molecules. Significance StatementThis study reveals that Phe84, a conserved aromatic residue in CD1d, may act as a ligand-responsive gate modulating F' groove accessibility. This conditional plasticity could enable binding of structurally diverse lipid antigens and appears conserved across CD1 isoforms. The mechanism parallels class I MHC, where gating residues regulate peptide presentation, suggesting an evolutionarily shared strategy for accommodating antigen diversity.

immunology↗

MAIT cells activate dendritic cells to promote T follicular helper cell differentiation and humoral immunity

Protective immune responses against respiratory pathogens, including influenza virus are initiated by the mucosal immune system. However, most licensed vaccines are administered parenterally and are largely ineffective at inducing mucosal immunity. The development of safe and effective mucosal vaccines has largely been hampered by the lack of a suitable mucosal adjuvant. In this study we explore a novel class of adjuvant that harness mucosal-associated invariant T (MAIT) cells. We show evidence that intranasal immunisation of MAIT cell agonists co-administered with protein, including haemagglutinin from influenza A virus induced potent humoral immunity and immunoglobulin (Ig)A production, which protected mice against infection. MAIT cell adjuvant activity was mediated by CD40L-dependent activation of dendritic cells and subsequent priming of CD4+ T follicular helper cells. In summary, we show that MAIT cells are promising vaccine targets that can be utilised as cellular adjuvants in mucosal vaccines.

immunology↗

The P5-ATPase ATP13A1 modulates MR1-mediated antigen presentation

The monomorphic antigen presenting molecule MHC-I-related protein 1 (MR1) presents small molecule metabolites to mucosal-associated invariant T (MAIT) cells. The MR1-MAIT cell axis has been implicated in a variety of infectious and non-communicable diseases and recent studies have begun to develop an understanding of the molecular mechanisms underlying this specialised antigen presentation pathway. Yet, the proteins regulating MR1 folding, loading, stability, and surface expression remain to be identified. Here, we performed a gene trap screen to discover novel modulators of MR1 surface expression through insertional mutagenesis of an MR1-overexpressing clone derived from the near-haploid human cell line HAP1 (HAP1.MR1). The most significant positive regulators identified included {beta}2-microglobulin, a known regulator of MR1 surface expression, and ATP13A1, a P5-ATPase in the endoplasmic reticulum (ER) with putative transporter function not previously associated with MR1-mediated antigen presentation. CRISPR/Cas9-mediated knock-out of ATP13A1 in both HAP1.MR1 and THP-1 cell lines revealed a profound reduction in MR1 protein levels and a concomitant functional defect specific to MR1-mediated antigen presentation. Collectively, these data are consistent with the ER-resident ATP13A1 as a key post-transcriptional determinant of MR1 surface expression.

immunology↗

Broad and strong memory CD4+ and CD8+ T cells induced by SARS-CoV-2 in UK convalescent COVID-19 patients.

COVID-19 is an ongoing global crisis in which the development of effective vaccines and therapeutics will depend critically on understanding the natural immunity to the virus, including the role of SARS-CoV-2-specific T cells. We have conducted a study of 42 patients following recovery from COVID-19, including 28 mild and 14 severe cases, comparing their T cell responses to those of 16 control donors. We assessed the immune memory of T cell responses using IFN{gamma} based assays with overlapping peptides spanning SARS-CoV-2 apart from ORF1. We found the breadth, magnitude and frequency of memory T cell responses from COVID-19 were significantly higher in severe compared to mild COVID-19 cases, and this effect was most marked in response to spike, membrane, and ORF3a proteins. Total and spike-specific T cell responses correlated with the anti-Spike, anti-Receptor Binding Domain (RBD) as well as anti-Nucleoprotein (NP) endpoint antibody titre (p<0.001, <0.001 and =0.002). We identified 39 separate peptides containing CD4+ and/or CD8+ epitopes, which strikingly included six immunodominant epitope clusters targeted by T cells in many donors, including 3 clusters in spike (recognised by 29%, 24%, 18% donors), two in the membrane protein (M, 32%, 47%) and one in the nucleoprotein (Np, 35%). CD8+ responses were further defined for their HLA restriction, including B*4001-restricted T cells showing central memory and effector memory phenotype. In mild cases, higher frequencies of multi-cytokine producing M- and NP-specific CD8+ T cells than spike-specific CD8+ T cells were observed. They furthermore showed a higher ratio of SARS-CoV-2-specific CD8+ to CD4+ T cell responses. Immunodominant epitope clusters and peptides containing T cell epitopes identified in this study will provide critical tools to study the role of virus-specific T cells in control and resolution of SARS-CoV-2 infections. The identification of T cell specificity and functionality associated with milder disease, highlights the potential importance of including non-spike proteins within future COVID-19 vaccine design.

immunology↗

Deep analysis of the USP18-dependent ISGylome and proteome unveils important roles for USP18 in tumour cell antigenicity and radiosensitivity

The deubiquitylating enzyme USP18 is a major negative regulator of the interferon (IFN) signalling cascade. IFN pathways contribute to resistance to conventional chemotherapy, radiotherapy, and immunotherapy and are often deregulated in cancer. USP18 is the predominant human protease that cleaves interferon-stimulated gene ISG15, a ubiquitin-like protein tightly regulated in the context of innate immunity, from its modified substrate proteins in vivo. In this study, using advanced proteomic techniques, we have expanded the USP18-dependent ISGylome and proteome in a chronic myeloid leukaemia (CML)-derived cell line (HAP1) treated with type I IFN. Novel ISGylation targets were characterised that modulate the sensing of innate ligands, antigen presentation and secretion of cytokines. Consequently, CML USP18-deficient cells are more antigenic, driving increased activation of cytotoxic T lymphocytes (CTLs) and are more susceptible to irradiation. Our results suggest USP18 as a pharmacological target in cancer immunotherapy and radiotherapy.

cancer biology↗

African Salmonella Typhimurium sequence type 313 lineage 2 evades MAIT cell recognition by overexpressing RibB

Mucosal-associated invariant T (MAIT) cells are a subset of innate T lymphocytes activated by bacteria that produce vitamin B2 metabolites. Mouse models of infection have demonstrated a role for MAIT cells in antimicrobial defence. However, proposed protective roles of MAIT cells in human infections remain unproven and clinical conditions associated with a selective absence of MAIT cells have not been identified. We report that typhoidal and non-typhoidal S. enterica strains generally activate MAIT cells. However, African invasive disease-associated multidrug-resistant S. Typhimurium sequence type 313 lineage 2 strains escape MAIT cell recognition through overexpression of ribB, a bacterial gene that encodes the 4-dihydroxy-2-butanone 4-phosphate synthase enzyme of the riboflavin biosynthetic pathway. This MAIT cell-specific phenotype did not extend to other innate lymphocytes. We propose that ribB overexpression is an evolved trait that facilitates evasion from immune recognition by MAIT cells and contributes to the invasive pathogenesis of S. Typhimurium sequence type 313 lineage 2 in vivo.

immunology↗