Biology subjects

Goodnow, C. C.

Publications and source records attributed to Goodnow, C. C..

10 recordsLinked to original sources

bioRxiv2026

Single-cell multi-omics maps clonal IEL expansion and epithelial remodelling in refractory coeliac disease

Background Refractory coeliac disease type 1 (RCD1) lacks defining molecular markers, and the immune and epithelial mechanisms sustaining intestinal injury remain poorly understood. Objective To define the clonal immune and epithelial states that distinguish RCD1 from active coeliac disease (ACD) and determine their spatial organisation in the duodenal mucosa. Design We integrated single-cell RNA sequencing, CITE-seq surface proteomics and paired T-cell receptor sequencing of duodenal immune and epithelial compartments from Healthy controls (n = 6), ACD (n = 7), RCD1 (n = 9) and RCD2 (n = 2), with spatial transcriptomics in a subset of biopsies. Results RCD1 showed widespread TCR{beta} and TCR{gamma}{delta} clonal expansion across multiple IEL states, extending beyond previously defined mutation-bearing aberrant clones. Distinct IEL populations converged on a shared programme of adaptive persistence, innate-like signalling, metabolic fitness and cytoskeletal remodelling; GZMK expression marked both clonally expanded and non-clonal disease-associated states. In parallel, RCD1 epithelium showed loss of mature absorptive cell states and expansion of stress-associated, immune-interacting and regenerative programmes. Transit-amplifying cells acquired differentiation and tissue-remodelling signatures, while enteroendocrine cells expanded and developed a sensory-neurosecretory programme involving TRPA1, TRPV1, vesicle trafficking and NEUROD1 regulon activity. Spatial transcriptomics localised regenerative and enteroendocrine-associated epithelial programmes adjacent to immune-visible epithelial regions and KLRK1/GZMK-expressing IEL-rich niches in refractory tissue. Conclusion RCD1 represents a distinct mucosal state characterised by coordinated clonal IEL adaptation and epithelial remodelling, rather than simple amplification of ACD, providing a cellular framework for persistent tissue injury and disease stratification.

immunology↗

bioRxiv2024

Self-reactive B cells traverse a perfect storm of somatic mutagenesis to cause a virus-induced autoimmune disease

The unexplained association between infection and autoimmune disease is strongest for hepatitis C virus-induced cryoglobulinemic vasculitis (HCV-CV). We traced the evolution of the pathogenic rheumatoid factor (RhF) autoantibodies in four HCV-CV patients by deep single cell multi-omic analysis, revealing three sources of B cell somatic mutation converged to drive accumulation of a large disease causing clone. A sensitive method for quantifying low affinity binding revealed three recurring heavy/light chain combinations created by V(D)J recombination bound self IgG but not viral E2 antigen. Whole genome sequencing revealed accumulation of thousands of somatic mutations, at levels comparable to CLL and normal memory B cells, but with 1-2 corresponding to driver mutations found recurrently in B cell leukemia/lymphoma. V(D)J hypermutation created autoantibodies with compromised solubility. In this virus-induced autoimmune disease, infection promotes a perfect storm of somatic mutagenesis in the descendants of a single B cell.

immunology↗

bioRxiv2023

DOCK2-deficiency causes defects in anti-viral T cell responses and poor control of herpes simplex virus infection

The expanding number of rare immunodeficiency syndromes offers an opportunity to understand key genes that support immune defence against infectious diseases. However, patients with these diseases are by definition rare. In addition, any analysis is complicated by treatments and co-morbid infections requiring the use of mouse models for detailed investigations. Here we develop a mouse model of DOCK2 immunodeficiency and demonstrate that these mice have delayed clearance of herpes simplex virus type 1 (HSV-1) infections. Further, we found that they have a critical, cell intrinsic role of DOCK2 in the clonal expansion of anti-viral CD8+ T cells despite normal early activation of these cells. Finally, while the major deficiency is in clonal expansion, the ability of primed and expanded DOCK2-deficient CD8+ T cells to protect against HSV-1-infection is also compromised. These results provide a contributing cause for the frequent and devastating viral infections seen in DOCK2-deficient patients and improve our understanding of anti-viral CD8+ T cell immunity.

immunology↗

bioRxiv2022

Integration of single-cell RNA and protein data identifies novel clinically-relevant lymphocyte phenotypes in breast cancers

Immune cells are critical determinants of solid tumour aetiology, but the diverse phenotypes of intra-tumoural immune cells remain incompletely characterised. We applied integrated single cell RNA sequencing (scRNA-Seq) and highly multiplexed protein epitope analysis to a cohort of breast cancer samples to resolve cell states within the tumour microenvironment. We reveal novel protein markers for resting and activated tumour infiltrating lymphocytes, and show that high expression of CD103 primarily marks exhausted CD8 rather than tissue resident CD8 T-cells in human breast cancers. We identify two distinct states of activated CD4+ T follicular helper (Tfh) cells. A population resembling conventional Tfh (cTfh) cells were localised primarily to lymphoid aggregates by spatial transcriptomics. In contrast, cancer associated Tfh (caTfh) cells expressing markers of tissue residency and exhaustion co-localized with cancer foci and signalled to macrophages. Importantly, increased caTfh : cTfh ratio associated with improved disease outcome and response to checkpoint immunotherapy.

immunology↗

bioRxiv2022

STAT3 gain-of-function mutations connect leukemia with autoimmune disease by pathological dysregulation of NKG2Dhi CD8 killer T cells

The association between cancer and autoimmune disease is unexplained, exemplified by T-cell large granular lymphocytic leukemia (T-LGL) where gain-of-function somatic mutations in STAT3 correlate with co-existing autoimmunity. To resolve whether these mutations are the cause or consequence of CD8 clonal expansions and autoimmunity, here we analyse patients with germline STAT3 GOF syndrome and mice with the T-LGL mutation STAT3K658Nor the most common germline mutation, STAT3T716M. STAT3 GOF mutations drove accumulation of effector CD8 T cell clones highly expressing the NKG2D receptor for MHC-I-related molecules expressed on stressed cells, and the genes for inflammatory/cytotoxic granzymes, perforin, interferon-{gamma} and Ccl5/Rantes. CD8 cells were essential to lethal disease in Stat3K658Nmice and their accumulation required NKG2D and the receptor for IL-15 and IL-2, IL2RB. These results demonstrate that STAT3 GOF mutations cause effector CD8 T cell oligoclonal accumulation and that these rogue T cells contribute to autoimmune pathology, supporting the hypothesis that somatic mutations in leukemia/lymphoma driver genes contribute to autoimmune disease. IN BRIEFLeukemia and autoimmune-associated STAT3 gain-of-function mutations dysregulate CD8 T cells to cause autoimmune pathology and oligoclonal expansion of cytotoxic killer CD8 T cells, that depend upon NKG2D and IL2RB receptors for signals displayed on stressed, damaged, infected, or mutated tissues.

immunology↗

bioRxiv2021

Overactive STAT3 drives accumulation of disease-associated CD21low B cells

Dysregulated STAT3 signalling is correlated with antibody-mediated autoimmunity and B- cell neoplasia, but its effect on B cells is underexplored. Here we address this in children with STAT3 gain-of-function (GOF) syndrome and in mice with STAT3T716M, the most common STAT3 GOF syndrome human mutation, or STAT3K658N, a dimerization interface mutation responsible for STAT3 GOF syndrome in two children. The main B cell consequence of overactive STAT3 was accumulation of CD19high CD21low atypical memory B cells in humans and of CD21low CD23low B cells in mice resembling age-associated B cells expressing T-bet, CD11c and plasma cell differentiation genes. Overactive STAT3 within B cells increased expression of many genes in the B cell receptor and T cell help pathways, increased the tolerogenic receptor CD22, but opposed B cell tolerance checkpoints and increased formation of T-bet+ B cells upon BCR and CD40 stimulation. These results reveal overactive STAT3 as a central driver of a key class of disease- associated B-lymphocytes in humans and mice.

immunology↗

bioRxiv2021

Dysregulation of PAX5 causes uncommitted B cell development and tumorigenesis in mice

PAX5 is the master transcription factor controlling B cell identity. In humans, mutations in PAX5 account for 30% of B cell acute lymphoblastic leukemia (B-ALL) cases. Investigating the causal effects of PAX5 mutations has however been difficult due to the premature lethality of Pax5-/- mice. Here we describe a novel mouse strain with a premature STOP mutation in Pax5 (Y351*) that produces a truncated protein and reduction in protein function, yet still allows for some B cell development to occur. A population of uncommitted and multipotent CD19+B220- B cells develops in the bone marrow of homozygous mice leading to the development of B-ALL. We show that the tumors frequently acquire secondary mutations in Jak3, and Ptpn11 highlighting key pathways interacting with PAX5 during malignant transformation. Analysis of the PAX5Y351* mice provide insight not only into the functional consequence of reduced PAX5 activity on B cell development and identity, but also provides an avenue in which to study PAX5-driven B-ALL in mice. One Sentence SummaryReduction in PAX5 function in mice induces the development of uncommitted B cells that have multipotent and malignant potential.

immunology↗

bioRxiv2020

Potent SARS-CoV-2 binding and neutralization through maturation of iconic SARS-CoV-1 antibodies

Antibodies against coronavirus spike protein potently protect against infection and disease, however it remains unclear if such protection can be extended to variant coronaviruses. This is exemplified by a set of iconic and well-characterized monoclonal antibodies developed after the 2003 SARS outbreak including mAbs m396, CR3022, CR3014 and 80R, which potently neutralize SARS-CoV-1, but not SARS-CoV-2. Here we explore antibody maturation strategies to change and broaden their specificity, enabling potent binding and neutralization of SARS-CoV-2. Using targeted mutagenesis as well as light chain shuffling on phage, we identified variants with considerably increased affinity and neutralization potential. The most potent antibody, derived from the NIH-developed mAb m396, neutralized live SARS-CoV-2 virus with a half-maximal inhibitory concentration (IC50) of 160 ng/ml. Intriguingly, while many of the matured clones maintained specificity of the parental antibody, new specificities were also observed, which was further confirmed by X-ray crystallography and cryo-electron microscopy, indicating that a limited set of antibodies can give rise to variants targeting diverse epitopes. Our findings open up over 15 years of antibody development efforts against SARS-CoV-1 to the SARS-CoV-2 field and outline general principles for the maturation of antibody specificity against emerging viruses.

immunology↗

bioRxiv2018

High-throughput targeted long-read single cell sequencing reveals the clonal and transcriptional landscape of lymphocytes

High-throughput single-cell RNA-Sequencing is a powerful technique for gene expression profiling of complex and heterogeneous cellular populations such as the immune system. However, these methods only provide short-read sequence from one end of a cDNA template, making them poorly suited to the investigation of gene-regulatory events such as mRNA splicing, adaptive immune responses or somatic genome evolution. To address this challenge, we have developed a method that combines targeted long-read sequencing with short-read based transcriptome profiling of barcoded single cell libraries generated by droplet-based partitioning. We use Repertoire And Gene Expression sequencing (RAGE-seq) to accurately characterize full-length T cell (TCR) and B cell (BCR) receptor sequences and transcriptional profiles of more than 7,138 lymphocytes sampled from the primary tumour and draining lymph node of a breast cancer patient. With this method we show that somatic mutation, alternate splicing and clonal evolution of T and B lymphocytes can be tracked across these tissue compartments. Our results demonstrate that RAGE-Seq is an accessible and cost-effective method for high-throughput deep single cell profiling, applicable to a wide range of biological challenges.

genomics↗

bioRxiv2017

B-cell receptor reconstruction from single-cell RNA-seq with VDJPuzzle

The B-cell receptor (BCR) performs essential functions for the adaptive immune system including recognition of pathogen-derived antigens. Cell-to-cell variability of BCR sequences due to V(D)J recombination and somatic hypermutation (SHM) necessitates single-cell characterization of BCR sequences. Single-cell RNA sequencing (scRNA-seq) presents the opportunity for simultaneous capture of the BCR sequence and transcriptomic signature for a detailed understanding of the dynamics of an immune response.\n\nWe developed VDJPuzzle 2.0, a bioinformatic tool that reconstructs productive, full-length B-cell receptor sequences of both heavy and light chains. VDJPuzzle successfully reconstructs BCRs from 98.3% (n=117) of human and 96.5% (n=200) from murine B cells. 92.0% of clonotypes and 90.3% of mutations were concordant with single-cell Sanger sequencing of the immunoglobulin chains. VDJPuzzle is available at https://bitbucket.org/kirbyvisp/vdjpuzzle2

bioinformatics↗