Biology subjects

James, E. A.

Publications and source records attributed to James, E. A..

6 recordsLinked to original sources

bioRxiv2026

Cathepsin K Mediates the Formation of Potential Rheumatoid Arthritis-Relevant Cis- and Trans-Spliced Peptides Compatible With HLA-DR4 Presentation

Rheumatoid arthritis (RA) is characterized by a loss of immunological tolerance to synovial self-proteins, yet the initial triggers generating novel neo-antigens remain incompletely defined. Here, we demonstrate that human cathepsin K (hCatK), a key cysteine protease driving joint degradation in RA, catalyzes covalent cis- and trans-splicing of peptides from major RA-associated self-proteins and foreign antigens, including type II collagen, fibrinogen, and SARS-CoV-2 Spike protein. Using high-resolution LC-MS/MS and database-assisted de novo sequencing, we identified over 90 unique spliced peptides. Splicing efficiency peaked at near-neutral pH (6.5-7.5), contrasting with classic hydrolytic profiles. Biochemical profiling revealed strong subsite selectivity, with a striking enrichment for small, aliphatic and/or hydroxyl-containing residues (Gly, Thr, Ser) at the P1 position. Furthermore, splicing preferentially targeted flexible, intrinsically disordered protein regions, with 81% of fibrinogen splicing events clustering within its C domain. In silico binding predictions for the RA-susceptibility allele HLA-DRB1*04:01 harboring the shared epitope revealed that numerous hCatK-generated spliced peptides exhibit predicted affinities exceeding those of established immunogenic and genomic sequences, uncovering protease-mediated transpeptidation as a novel post-translational modification capable of generating potent MHC class II autoantigens in RA.

biochemistry↗

bioRxiv2025

Long non-coding RNAs as a novel source of beta cell autoantigens in type 1 diabetes

Aims/hypothesisGenome-wide association studies increasingly highlight the role of non-coding regions in complex diseases such as type 1 diabetes, pointing, in particular, to long non-coding RNAs (lncRNAs) as potentially active molecular players. Emerging evidence suggests that lncRNAs may encode (small) peptides with the potential to modulate cellular processes, including immune responses. In this study, we have investigated whether these (micro)peptides translated from lncRNAs can act as neoantigens capable of activating autoreactive CD4T cells in individuals with type 1 diabetes. MethodsWe have integrated transcriptomic, proteomic and in silico data to identify a subset of lncRNAs with genuine peptide-coding potential in basal or inflamed-condition beta cells. We further assessed the suitability of those candidates to encode (micro)peptides by sequence analysis and in vitro experimentation techniques. Finally, we have predicted HLA-II epitopes within the micropeptide candidates and evaluated their immunogenicity by analyzing T cell activation responses in peripheral blood mononuclear cells from HLA matching individuals with type 1 diabetes. ResultsBy merging ribosome-bound RNA sequencing with nascent peptide mass spectrometry, we identified a total of 30 lncRNAs with potential coding capacity. We verified the peptide-coding ORF translation for three lncRNA candidates: UXT-AS1, RAPGEF4-AS1 and ENSG00000227066. After predicting potential type 1 diabetes risk-associated HLA-DRB1*03:01 and HLA-DRB1*04:01-binding epitopes within translated lncRNA (micro)peptides, we identified several peptides that elicited CD4+ T cell activation. Furthermore, several epitopes elicited T cell activation in multiple donors with type 1 diabetes. T cell lines were isolated and studied to confirm responses restricted by type 1 diabetes risk HLA alleles. ConclusionsThese results reveal a novel class of immunogenic (micro)peptides derived from lncRNAs, supporting their potential role in the autoimmune response that underlies type 1 diabetes. Our findings open new perspectives on the contribution of non-coding genomic elements to autoimmunity and highlight the need to further investigate lncRNA-encoded (micro)peptides as possible targets for future immunotherapies. RESEARCH IN CONTEXTO_ST_ABSWhat is already known about this subject?C_ST_ABSO_LIShort proteins can sometimes be translated from lncRNAs. C_LIO_LIType 1 diabetes neoepitopes arising from non-native forms of proteins and their post translational modifications have been described. C_LI What is the key question?O_LICan lncRNA-encoded peptides be recognized by the human immune system and contribute to type 1 diabetes development? C_LI What are the new findings?O_LIWe have identified three candidate lncRNAs with peptide coding capacity. C_LIO_LIEpitopes derived from these peptides drive T cell activation in PBMCs of individuals with type 1 diabetes. C_LIO_LIPresentation of some of these peptides are restricted to type 1 diabetes risk-associated HLA-DRB1*03:01 and *04:01 molecules. C_LI How might this impact on clinical practice in the foreseeable future?O_LIDiscovery of lncRNA-derived peptide reactive T cells might serve as a diagnostic or stratification indicator in the early stages of disease development, aiding the development of more personalized treatments and unlocking a new array of therapeutic targets. C_LI

immunology↗

bioRxiv2025

Senescent Activated Naive B Cells Promote Anti-Citrullinated Antigen T Cell Responses and the Transition to Clinical Rheumatoid Arthritis

Rheumatoid arthritis (RA) is a chronic autoimmune disease marked by joint and systemic inflammation. Anti-citrullinated protein antibodies (ACPAs) define an at-risk stage that precedes clinically apparent inflammatory arthritis (clinical RA) onset, yet the molecular mechanisms driving progression remain poorly understood. Here, we applied single-cell multi-omics to profile B cells longitudinally collected from ACPA individuals who either convert to clinical RA (Converters) or do not (Nonconverters). We identified a striking expansion of CXCR5CD69 activated naive B cells (aNAVs) uniquely in Converters prior to clinical RA. These aNAVs exhibited a pro-inflammatory, senescent transcriptional program and persist through to clinical RA. In Converters, aNAVs expressed polyreactive, autoreactive IgM with distinctive V-J gene rearrangements that dominate the BCR repertoire. Furthermore, in Converters most IgM aNAVs were developmentally arrested in the peripheral blood, while a subset undergoes class switching and follows divergent somatic hypermutation trajectories. Mechanistically, aNAVs infiltrated RA synovium and served as potent antigen presenting cells to activate both anti-citrullinated antigen CD4 and CD8 T cells in an HLA-dependent manner. Chronic exposure to citrullinated antigens and CpG synergistically drove aNAV activation and senescence. These findings establish a mechanistic link between naive B cell senescence and clinical RA development in ACPA+ individuals, providing a rationale for therapeutically targeting aNAV B cells for the prevention of RA. Graphic abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=182 SRC="FIGDIR/small/682430v1_ufig1.gif" ALT="Figure 1"> View larger version (85K): org.highwire.dtl.DTLVardef@153b185org.highwire.dtl.DTLVardef@1aba6eeorg.highwire.dtl.DTLVardef@5c886eorg.highwire.dtl.DTLVardef@10109a3_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2022

Dual IgA/IgG family autoantibodies from individuals at-risk for rheumatoid arthritis identify an arthritogenic strain of Subdoligranulum

The mucosal origins hypothesis of rheumatoid arthritis (RA) proposes a central role for mucosal immune responses in the initiation and/or perpetuation of the systemic autoimmunity that occurs with disease. However, the connection between the mucosa and systemic autoimmunity in RA remains unclear. Using dual IgA/IgG family plasmablast-derived monoclonal autoantibodies obtained from peripheral blood of individuals at-risk for RA, we identified cross-reactivity between RA-relevant autoantigens and bacterial taxa in the closely related families Lachnospiraceae and Ruminococcaceae. After generating bacterial isolates within Lachnospiraceae/Ruminococcaceae genus Subdoligranulum from the feces of an individual, we confirmed monoclonal antibody binding as well as MHC class II dependent CD4+ T cell activation in RA cases compared to controls. Additionally, when Subdoligranulum isolate 7 but not isolate 1 colonized germ-free mice, it stimulated Th17 cell expansion, serum RA-relevant IgG autoantibodies, and joint swelling reminiscent of early RA with histopathology characterized by antibody deposition and complement activation. Systemic immune responses were likely due to the generation of colon isolated lymphoid follicles (ILFs) driving increased fecal and serum IgA by isolate 7, as B and T cell depletion not only halted intestinal immune responses but also eliminated detectable clinical disease. In aggregate, these findings demonstrate one mucosal mechanism in RA through which an intestinal strain of bacteria can drive systemic autoantibody generation and joint-centered antibody deposition and immune activation. One-sentence summarySubdoligranulum spp. targeted by rheumatoid arthritis (RA)-relevant autoantibodies activate T cells in individuals with RA, and in mice stimulate autoantibodies and joint swelling associated with antibody deposition and complement activation.

immunology↗

bioRxiv2021

Carbonyl Post-Translational Modification Associated with Early Onset Type 1 Diabetes Autoimmunity

Inflammation and oxidative stress in pancreatic islets amplify the appearance of various post-translational modifications (PTMs) to self-proteins. Herein, we identified a select group of carbonylated islet proteins arising before the onset of hyperglycemia in non-obese diabetic (NOD) mice. Of particular interest, we identified carbonyl modification of the prolyl-4-hydroxylase beta subunit (P4Hb) that is responsible for proinsulin folding and trafficking as an autoantigen in both human and murine type 1 diabetes. We found the carbonylated-P4Hb is amplified in stressed islets coincident with decreased glucose-stimulated insulin secretion and altered proinsulin to insulin ratios. Moreover, circulating autoantibodies against P4Hb were detected in prediabetic NOD mice and in early human type 1 diabetes prior to the onset of anti-insulin autoimmunity. Our studies provide mechanistic insight into the pathways of proinsulin metabolism and those creating autoantigenic forms of insulin in type 1 diabetes.

immunology↗

bioRxiv2021

Citrullination of glucokinase linked to autoimmune diabetes

Inflammation, including reactive oxygen species and inflammatory cytokines in tissue microenvironments amplify the appearance of various post-translational modifications (PTMs) of self-proteins. Previously, a number of PTMs have been identified as autoimmune biomarkers in the initiation and progression of Type 1 diabetes (T1D). Herein, we have identified the citrullination of glucokinase (GK) as a result of inflammation, triggering autoimmunity and affecting its biological functions. Glucokinase is predominantly expressed in hepatocytes to regulate glycogen synthesis, and in pancreatic beta cells, where it acts as a glucose sensor to initiate glycolysis and insulin signaling. Herein, we demonstrate that glucokinase is citrullinated in inflamed non-obese diabetic (NOD) islets as well as in human GK. Autoantibodies against both native and citrullinated GK arise in both spontaneous human T1D and murine models. Likewise, autoreactive CD4+ T cells to both native and citrullinated glucokinase epitopes are present in the circulation of T1D patients. Finally, citrullination alters GK biologic activity and suppresses glucose-stimulated insulin secretion. Our studies define glucokinase as a T1D biomarker, providing new insights into altering glucose metabolism, creating neoautoantigens, and better define the broad impact of PTMs on the tissue pathology of T1D.

immunology↗