Publications by authors named "Richard A Flavell"

693 Publications

mA demethylase ALKBH5 controls CD4 T cell pathogenicity and promotes autoimmunity.

Sci Adv 2021 Jun 16;7(25). Epub 2021 Jun 16.

Shanghai Institute of Immunology, State Key Laboratory of Oncogenes and Related Genes, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China.

N-methyladenosine (mA) modification is dynamically regulated by "writer" and "eraser" enzymes. mA "writers" have been shown to ensure the homeostasis of CD4 T cells, but the "erasers" functioning in T cells is poorly understood. Here, we reported that mA eraser AlkB homolog 5 (ALKBH5), but not FTO, maintains the ability of naïve CD4 T cells to induce adoptive transfer colitis. In addition, T cell-specific ablation of ALKBH5 confers protection against experimental autoimmune encephalomyelitis. During the induced neuroinflammation, ALKBH5 deficiency increased mA modification on interferon-γ and C-X-C motif chemokine ligand 2 messenger RNA (mRNA), thus decreasing their mRNA stability and protein expression in CD4 T cells. These modifications resulted in attenuated CD4 T cell responses and diminished recruitment of neutrophils into the central nervous system. Our findings reveal an unexpected specific role of ALKBH5 as an mA eraser in controlling the pathogenicity of CD4 T cells during autoimmunity.
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http://dx.doi.org/10.1126/sciadv.abg0470DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8208713PMC
June 2021

Modulating HIV-1 envelope glycoprotein conformation to decrease the HIV-1 reservoir.

Cell Host Microbe 2021 Jun 20;29(6):904-916.e6. Epub 2021 May 20.

Section of Infectious Diseases, Department of Internal Medicine, Yale University School of Medicine, New Haven, CT, USA. Electronic address:

Small CD4-mimetic compounds (CD4mc) sensitize HIV-1-infected cells to antibody-dependent cellular cytotoxicity (ADCC) by facilitating antibody recognition of epitopes that are otherwise occluded on the unliganded viral envelope (Env). Combining CD4mc with two families of CD4-induced (CD4i) antibodies, which are frequently found in plasma of HIV-1-infected individuals, stabilizes Env in a conformation that is vulnerable to ADCC. We employed new-generation SRG-15 humanized mice, supporting natural killer (NK) cell and Fc-effector functions to demonstrate that brief treatment with CD4mc and CD4i-Abs significantly decreases HIV-1 replication, the virus reservoir and viral rebound after ART interruption. These effects required Fc-effector functions and NK cells, highlighting the importance of ADCC. Viral rebound was also suppressed in HIV-1+-donor cell-derived humanized mice supplemented with autologous HIV-1+-donor-derived plasma and CD4mc. These results indicate that CD4mc could have therapeutic utility in infected individuals for decreasing the size of the HIV-1 reservoir and/or achieving a functional cure.
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http://dx.doi.org/10.1016/j.chom.2021.04.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8214472PMC
June 2021

Detection of differentially abundant cell subpopulations in scRNA-seq data.

Proc Natl Acad Sci U S A 2021 06;118(22)

Department of Pathology, Yale University, New Haven, CT 06511.

Comprehensive and accurate comparisons of transcriptomic distributions of cells from samples taken from two different biological states, such as healthy versus diseased individuals, are an emerging challenge in single-cell RNA sequencing (scRNA-seq) analysis. Current methods for detecting differentially abundant (DA) subpopulations between samples rely heavily on initial clustering of all cells in both samples. Often, this clustering step is inadequate since the DA subpopulations may not align with a clear cluster structure, and important differences between the two biological states can be missed. Here, we introduce DA-seq, a targeted approach for identifying DA subpopulations not restricted to clusters. DA-seq is a multiscale method that quantifies a local DA measure for each cell, which is computed from its nearest neighboring cells across a range of values. Based on this measure, DA-seq delineates contiguous significant DA subpopulations in the transcriptomic space. We apply DA-seq to several scRNA-seq datasets and highlight its improved ability to detect differences between distinct phenotypes in severe versus mildly ill COVID-19 patients, melanomas subjected to immune checkpoint therapy comparing responders to nonresponders, embryonic development at two time points, and young versus aging brain tissue. DA-seq enabled us to detect differences between these phenotypes. Importantly, we find that DA-seq not only recovers the DA cell types as discovered in the original studies but also reveals additional DA subpopulations that were not described before. Analysis of these subpopulations yields biological insights that would otherwise be undetected using conventional computational approaches.
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http://dx.doi.org/10.1073/pnas.2100293118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8179149PMC
June 2021

Pooled CRISPR screening identifies mA as a positive regulator of macrophage activation.

Sci Adv 2021 Apr 28;7(18). Epub 2021 Apr 28.

Shanghai Institute of Immunology, State Key Laboratory of Oncogenes and Related Genes, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China.

mA RNA modification is implicated in multiple cellular responses. However, its function in the innate immune cells is poorly understood. Here, we identified major mA "writers" as the top candidate genes regulating macrophage activation by LPS in an RNA binding protein focused CRISPR screening. We have confirmed that deficient macrophages exhibited reduced TNF-α production upon LPS stimulation in vitro. Consistently, ;Cre mice displayed increased susceptibility to bacterial infection and showed faster tumor growth. Mechanistically, the transcripts of the gene encoding a negative regulator of TLR4 signaling were highly decorated by mA modification. METTL3 deficiency led to the loss of mA modification on mRNA and slowed down its degradation, resulting in a higher level of IRAKM, which ultimately suppressed TLR signaling-mediated macrophage activation. Our findings demonstrate a previously unknown role for METTL3-mediated mA modification in innate immune responses and implicate the mA machinery as a potential cancer immunotherapy target.
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http://dx.doi.org/10.1126/sciadv.abd4742DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8081357PMC
April 2021

A humanized mouse model of chronic COVID-19 to evaluate disease mechanisms and treatment options.

Res Sq 2021 Mar 17. Epub 2021 Mar 17.

Coronavirus-associated acute respiratory disease, called coronavirus disease 2019 (COVID-19) is an infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). More than 90 million people have been infected with SARS-CoV-2 and more than 2 million people have died of complications due to COVID-19 worldwide. COVID-19, in its severe form, presents with an uncontrolled, hyperactive immune response and severe immunological injury or organ damage that accounts for morbidity and mortality. Even in the absence of complications, COVID-19 can last for several months with lingering effects of an overactive immune system. Dysregulated myeloid and lymphocyte compartments have been implicated in lung immunopathology. Currently, there are limited clinically-tested treatments of COVID-19 with disparities in the apparent efficacy in patients. Accurate model systems are essential to rapidly evaluate promising discoveries but most currently available in mice, ferrets and hamsters do not recapitulate sustained immunopathology described in COVID19 patients. Here, we present a comprehensively humanized mouse COVID-19 model that faithfully recapitulates the innate and adaptive human immune responses during infection with SARS-CoV-2 by adapting recombinant adeno-associated virus (AAV)-driven gene therapy to deliver human ACE2 to the lungs of MISTRG6 mice. Our unique model allows for the first time the study of chronic disease due to infection with SARS-CoV-2 in the context of patient-derived antibodies to characterize in real time the potential culprits of the observed human driving immunopathology; most importantly this model provides a live view into the aberrant macrophage response that is thought to be the effector of disease morbidity and ARDS in patients. Application of therapeutics such as patient-derived antibodies and steroids to our model allowed separation of the two aspects of the immune response, infectious viral clearance and immunopathology. Inflammatory cells seeded early in infection drove immune-patholgy later, but this very same early anti-viral response was also crucial to contain infection.
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http://dx.doi.org/10.21203/rs.3.rs-279341/v1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7987100PMC
March 2021

Cerebellar Kv3.3 potassium channels activate TANK-binding kinase 1 to regulate trafficking of the cell survival protein Hax-1.

Nat Commun 2021 03 19;12(1):1731. Epub 2021 Mar 19.

Department of Pharmacology, Yale University School of Medicine, New Haven, CT, USA.

Mutations in KCNC3, which encodes the Kv3.3 potassium channel, cause degeneration of the cerebellum, but exactly how the activity of an ion channel is linked to the survival of cerebellar neurons is not understood. Here, we report that Kv3.3 channels bind and stimulate Tank Binding Kinase 1 (TBK1), an enzyme that controls trafficking of membrane proteins into multivesicular bodies, and that this stimulation is greatly increased by a disease-causing Kv3.3 mutation. TBK1 activity is required for the binding of Kv3.3 to its auxiliary subunit Hax-1, which prevents channel inactivation with depolarization. Hax-1 is also an anti-apoptotic protein required for survival of cerebellar neurons. Overactivation of TBK1 by the mutant channel leads to the loss of Hax-1 by its accumulation in multivesicular bodies and lysosomes, and also stimulates exosome release from neurons. This process is coupled to activation of caspases and increased cell death. Our studies indicate that Kv3.3 channels are directly coupled to TBK1-dependent biochemical pathways that determine the trafficking of cellular constituents and neuronal survival.
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http://dx.doi.org/10.1038/s41467-021-22003-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7979925PMC
March 2021

Combined liver-cytokine humanization comes to the rescue of circulating human red blood cells.

Science 2021 03;371(6533):1019-1025

Department of Immunobiology, Yale University School of Medicine, New Haven, CT, USA.

In vivo models that recapitulate human erythropoiesis with persistence of circulating red blood cells (RBCs) have remained elusive. We report an immunodeficient murine model in which combined human liver and cytokine humanization confer enhanced human erythropoiesis and RBC survival in the circulation. We deleted the fumarylacetoacetate hydrolase () gene in MISTRG mice expressing several human cytokines in place of their murine counterparts. Liver humanization by intrasplenic injection of human hepatocytes (huHep) eliminated murine complement C3 and reduced murine Kupffer cell density. Engraftment of human sickle cell disease (SCD)-derived hematopoietic stem cells in huHepMISTRG mice resulted in vaso-occlusion that replicated acute SCD pathology. Combined liver-cytokine-humanized mice will facilitate the study of diseases afflicting RBCs, including bone marrow failure, hemoglobinopathies, and malaria, and also preclinical testing of therapies.
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http://dx.doi.org/10.1126/science.abe2485DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8292008PMC
March 2021

MAP3K2-regulated intestinal stromal cells define a distinct stem cell niche.

Nature 2021 Apr 3;592(7855):606-610. Epub 2021 Mar 3.

Shanghai Institute of Immunology, Department of Immunology and Microbiology, Shanghai Jiao Tong University School of Medicine, Shanghai, China.

Intestinal stromal cells are known to modulate the propagation and differentiation of intestinal stem cells. However, the precise cellular and molecular mechanisms by which this diverse stromal cell population maintains tissue homeostasis and repair are poorly understood. Here we describe a subset of intestinal stromal cells, named MAP3K2-regulated intestinal stromal cells (MRISCs), and show that they are the primary cellular source of the WNT agonist R-spondin 1 following intestinal injury in mice. MRISCs, which are epigenetically and transcriptomically distinct from subsets of intestinal stromal cells that have previously been reported, are strategically localized at the bases of colon crypts, and function to maintain LGR5 intestinal stem cells and protect against acute intestinal damage through enhanced R-spondin 1 production. Mechanistically, this MAP3K2 specific function is mediated by a previously unknown reactive oxygen species (ROS)-MAP3K2-ERK5-KLF2 axis to enhance production of R-spondin 1. Our results identify MRISCs as a key component of an intestinal stem cell niche that specifically depends on MAP3K2 to augment WNT signalling for the regeneration of damaged intestine.
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http://dx.doi.org/10.1038/s41586-021-03283-yDOI Listing
April 2021

JUN Amino-Terminal Kinase 1 Signaling in the Proximal Tubule Causes Cell Death and Acute Renal Failure in Rat and Mouse Models of Renal Ischemia/Reperfusion Injury.

Am J Pathol 2021 05 16;191(5):817-828. Epub 2021 Feb 16.

Department of Nephrology, Monash Medical Centre, Clayton, Victoria, Australia; Centre for Inflammatory Diseases, Monash University, Clayton, Victoria, Australia.

Activation of the JUN amino-terminal kinase (JNK) pathway is prominent in most forms of acute and progressive tubulointerstitial damage, including acute renal ischemia/reperfusion injury (IRI). Two forms of JNK, JNK1 and JNK2, are expressed in the kidney. Systemic administration of pan-JNK inhibitors suppresses renal IRI; however, the contribution of JNK1 versus JNK2, and the specific role of JNK activation in the proximal tubule in IRI, remains unknown. These questions were addressed in rat and mouse models of acute bilateral renal IRI. Administration of the JNK inhibitor, CC-930, substantially reduced the severity of renal failure, tubular damage, and inflammation at 24 hours in a rat IRI model. Additionally, Jnk1 mice, but not Jnk2 mice, were shown to be significantly protected against acute renal failure, tubular damage, and inflammation in the IRI model. Furthermore, mice with conditional Jnk1 deletion in the proximal tubule also showed considerable protection from IRI-induced renal failure, tubular damage, and inflammation. Finally, primary cultures of Jnk1, but not Jnk2, tubular epithelial cells were protected from oxidant-induced cell death, in association with preventing phosphorylation of proteins (receptor interacting serine/threonine kinase 3 and mixed lineage kinase domain-like pseudokinase) in the necroptosis pathway. In conclusion, JNK1, but not JNK2, plays a specific role in IRI-induced cell death in the proximal tubule, leading to acute renal failure.
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http://dx.doi.org/10.1016/j.ajpath.2021.02.004DOI Listing
May 2021

Role of MBD3-SOX2 axis in residual myeloma following pomalidomide.

Leukemia 2021 Feb 18. Epub 2021 Feb 18.

Winship Cancer Institute, Emory University, Atlanta, GA, USA.

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http://dx.doi.org/10.1038/s41375-021-01145-0DOI Listing
February 2021

Potential intestinal infection and faecal-oral transmission of SARS-CoV-2.

Nat Rev Gastroenterol Hepatol 2021 04 15;18(4):269-283. Epub 2021 Feb 15.

Department of Digestive Disease, The First Affiliated Hospital of USTC, Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei, China.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread to more than 200 countries and regions globally. SARS-CoV-2 is thought to spread mainly through respiratory droplets and close contact. However, reports have shown that a notable proportion of patients with coronavirus disease 2019 (COVID-19) develop gastrointestinal symptoms and nearly half of patients confirmed to have COVID-19 have shown detectable SARS-CoV-2 RNA in their faecal samples. Moreover, SARS-CoV-2 infection reportedly alters intestinal microbiota, which correlated with the expression of inflammatory factors. Furthermore, multiple in vitro and in vivo animal studies have provided direct evidence of intestinal infection by SARS-CoV-2. These lines of evidence highlight the nature of SARS-CoV-2 gastrointestinal infection and its potential faecal-oral transmission. Here, we summarize the current findings on the gastrointestinal manifestations of COVID-19 and its possible mechanisms. We also discuss how SARS-CoV-2 gastrointestinal infection might occur and the current evidence and future studies needed to establish the occurrence of faecal-oral transmission.
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http://dx.doi.org/10.1038/s41575-021-00416-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7883337PMC
April 2021

Skin-resident innate lymphoid cells converge on a pathogenic effector state.

Nature 2021 04 3;592(7852):128-132. Epub 2021 Feb 3.

Department of Medicine, Sandler Asthma Research Center, University of California San Francisco, San Francisco, CA, USA.

Tissue-resident innate lymphoid cells (ILCs) help sustain barrier function and respond to local signals. ILCs are traditionally classified as ILC1, ILC2 or ILC3 on the basis of their expression of specific transcription factors and cytokines. In the skin, disease-specific production of ILC3-associated cytokines interleukin (IL)-17 and IL-22 in response to IL-23 signalling contributes to dermal inflammation in psoriasis. However, it is not known whether this response is initiated by pre-committed ILCs or by cell-state transitions. Here we show that the induction of psoriasis in mice by IL-23 or imiquimod reconfigures a spectrum of skin ILCs, which converge on a pathogenic ILC3-like state. Tissue-resident ILCs were necessary and sufficient, in the absence of circulatory ILCs, to drive pathology. Single-cell RNA-sequencing (scRNA-seq) profiles of skin ILCs along a time course of psoriatic inflammation formed a dense transcriptional continuum-even at steady state-reflecting fluid ILC states, including a naive or quiescent-like state and an ILC2 effector state. Upon disease induction, the continuum shifted rapidly to span a mixed, ILC3-like subset also expressing cytokines characteristic of ILC2s, which we inferred as arising through multiple trajectories. We confirmed the transition potential of quiescent-like and ILC2 states using in vitro experiments, single-cell assay for transposase-accessible chromatin using sequencing (scATAC-seq) and in vivo fate mapping. Our results highlight the range and flexibility of skin ILC responses, suggesting that immune activities primed in healthy tissues dynamically adapt to provocations and, left unchecked, drive pathological remodelling.
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http://dx.doi.org/10.1038/s41586-021-03188-wDOI Listing
April 2021

Presynaptic Kv3 channels are required for fast and slow endocytosis of synaptic vesicles.

Neuron 2021 03 27;109(6):938-946.e5. Epub 2021 Jan 27.

National Institute of Neurological Disorders and Stroke, 35 Convent Dr., Bethesda, MD 20892, USA. Electronic address:

Since their discovery decades ago, the primary physiological and pathological effects of potassium channels have been attributed to their ion conductance, which sets membrane potential and repolarizes action potentials. For example, Kv3 family channels regulate neurotransmitter release by repolarizing action potentials. Here we report a surprising but crucial function independent of potassium conductance: by organizing the F-actin cytoskeleton in mouse nerve terminals, the Kv3.3 protein facilitates slow endocytosis, rapid endocytosis, vesicle mobilization to the readily releasable pool, and recovery of synaptic depression during repetitive firing. A channel mutation that causes spinocerebellar ataxia inhibits endocytosis, vesicle mobilization, and synaptic transmission during repetitive firing by disrupting the ability of the channel to nucleate F-actin. These results unmask novel functions of potassium channels in endocytosis and vesicle mobilization crucial for sustaining synaptic transmission during repetitive firing. Potassium channel mutations that impair these "non-conducting" functions may thus contribute to generation of diverse neurological disorders.
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http://dx.doi.org/10.1016/j.neuron.2021.01.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7979485PMC
March 2021

The RNA helicase Dhx15 mediates Wnt-induced antimicrobial protein expression in Paneth cells.

Proc Natl Acad Sci U S A 2021 01;118(4)

Department of Digestive Disease, The First Affiliated Hospital of University of Science and Technology of China, Division of Life Sciences and Medicine, University of Science and Technology of China, 230001 Hefei, China;

RNA helicases play roles in various essential biological processes such as RNA splicing and editing. Recent in vitro studies show that RNA helicases are involved in immune responses toward viruses, serving as viral RNA sensors or immune signaling adaptors. However, there is still a lack of in vivo data to support the tissue- or cell-specific function of RNA helicases owing to the lethality of mice with complete knockout of RNA helicases; further, there is a lack of evidence about the antibacterial role of helicases. Here, we investigated the in vivo role of Dhx15 in intestinal antibacterial responses by generating mice that were intestinal epithelial cell (IEC)-specific deficient for Dhx15 (Dhx15 f/f Villin1-cre, Dhx15). These mice are susceptible to infection with enteric bacteria (), owing to impaired α-defensin production by Paneth cells. Moreover, mice with Paneth cell-specific depletion of Dhx15 (Dhx15 f/f Defensinα6-cre, Dhx15) are more susceptible to DSS (dextran sodium sulfate)-induced colitis, which phenocopy Dhx15 mice, due to the dysbiosis of the intestinal microbiota. In humans, reduced protein levels of Dhx15 are found in ulcerative colitis (UC) patients. Taken together, our findings identify a key regulator of Wnt-induced α-defensins in Paneth cells and offer insights into its role in the antimicrobial response as well as intestinal inflammation.
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http://dx.doi.org/10.1073/pnas.2017432118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7848544PMC
January 2021

Enoxacin upregulates microRNA biogenesis and downregulates cytotoxic CD8 T cell function in autoimmune cholangitis.

Hepatology 2021 Jan 19. Epub 2021 Jan 19.

Division of Immunology, Allergy and Rheumatology, University of Cincinnati College of Medicine, Cincinnati, Ohio, 45267, USA.

Background And Aims: Primary biliary cholangitis (PBC) is a prototypical organ specific autoimmune disease, mediated by autoreactive T cell attack and destruction of cholangiocytes. Despite the clear role of autoimmunity in PBC, immune-directed therapies have failed to halt PBC, including biologic therapies effective in other autoimmune disease. MicroRNA (miRNA) dysregulation is implicated in the pathogenesis (PBC). In the dominant negative TGF-β receptor (dnTGFβRII) mouse model of PBC, autoreactive CD8 T cells play a major pathogenic role, and demonstrate a striking pattern of miRNA downregulation. Enoxacin is a small molecule fluoroquinolone that enhances miRNA biogenesis, partly by stabilizing the interaction of trans-activation response RNA binding protein (TRBP) with Argonaute2 (Ago2).

Approach And Results: We hypothesized that correcting aberrant T cell miRNA expression with enoxacin in dnTGFβRII mice could modulate autoreactive T cell function and prevent PBC. Here we show that liver-infiltrating dnTGFβRII CD8 T cells have significantly decreased levels of the miRNA biogenesis molecules P4ha1 and Ago2 along with significantly increased levels of granzyme B and perforin. Enoxacin treatment significantly upregulated miRNAs in dnTGFβRII CD8 T cells and effectively treated autoimmune cholangitis in dnTGFβRII mice. Enoxacin treatment directly altered T cells both ex vivo and in vitro, resulting in altered memory subset numbers, decreased proliferation and decreased IFN-γ production. Enoxacin significantly decreased CD8 T cell expression of the transcription factor, Runx3, and significantly decreased perforin expression at both the mRNA and protein levels.

Conclusions: Enoxacin increases miRNA expression in dnTGFβRII CD8 T cells, reduces CD8 T cell pathogenicity, and effectively halted progression of autoimmune biliary disease. Targeting the miRNA pathway is a novel therapeutic approach to autoimmunity that corrects pathological miRNA abnormalities in autoreactive T cells.
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http://dx.doi.org/10.1002/hep.31724DOI Listing
January 2021

An in vivo screen of noncoding loci reveals that is a gatekeeper of an Ikaros-dependent checkpoint during haematopoiesis.

Proc Natl Acad Sci U S A 2021 01;118(3)

Howard Hughes Medical Institute, New Haven, CT 06520;

Haematopoiesis relies on tightly controlled gene expression patterns as development proceeds through a series of progenitors. While the regulation of hematopoietic development has been well studied, the role of noncoding elements in this critical process is a developing field. In particular, the discovery of new regulators of lymphopoiesis could have important implications for our understanding of the adaptive immune system and disease. Here we elucidate how a noncoding element is capable of regulating a broadly expressed transcription factor, Ikaros, in a lymphoid lineage-specific manner, such that it imbues Ikaros with the ability to specify the lymphoid lineage over alternate fates. Deletion of the locus, which is proximal to Ikaros, led to a severe reduction in early lymphoid progenitors, exerting control over the earliest fate decisions during lymphoid lineage commitment. locus deletion led to alterations in Ikaros isoform expression and a significant reduction in Ikaros protein. The locus may function through direct DNA interaction as Hi-C analysis demonstrated an interaction between the two loci. Finally, we identify an Ikaros-regulated erythroid-lymphoid checkpoint that is governed by in a lymphoid-lineage-specific manner. appears to act as a gatekeeper of Ikaros's broad lineage-specifying functions, selectively stabilizing Ikaros activity in the lymphoid lineage and permitting diversion to the erythroid fate in its absence. These findings represent a key illustration of how a transcription factor with broad lineage expression must work in concert with noncoding elements to orchestrate hematopoietic lineage commitment.
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http://dx.doi.org/10.1073/pnas.1918062118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7826330PMC
January 2021

Immunoglobulin A Targets a Unique Subset of the Microbiota in Inflammatory Bowel Disease.

Cell Host Microbe 2021 01 1;29(1):83-93.e3. Epub 2021 Jan 1.

Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA. Electronic address:

The immunopathogenesis of inflammatory bowel disease (IBD) has been attributed to a combination of host genetics and intestinal dysbiosis. Previous work in a small cohort of IBD patients suggested that pro-inflammatory bacterial taxa are highly coated with secretory immunoglobulin IgA. Using bacterial fluorescence-activated cell sorting coupled with 16S rRNA gene sequencing (IgA-SEQ), we profiled IgA coating of intestinal microbiota in a large cohort of IBD patients and identified bacteria associated with disease and treatment. Forty-three bacterial taxa displayed significantly higher IgA coating in IBD compared with controls, including 8 taxa exhibiting differential IgA coating but similar relative abundance. Patients treated with anti-TNF-α therapies exhibited dramatically altered microbiota-specific IgA responses compared with controls. Furthermore, increased IgA coating of Oscillospira was associated with a delay in time to surgery. These results demonstrate that investigating IgA responses to microbiota can uncover potential disease-modifying taxa and reveal improved biomarkers of clinical course in IBD.
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http://dx.doi.org/10.1016/j.chom.2020.12.003DOI Listing
January 2021

Treatment of granuloma annulare and suppression of proinflammatory cytokine activity with tofacitinib.

J Allergy Clin Immunol 2021 May 11;147(5):1795-1809. Epub 2020 Dec 11.

Department of Dermatology, Yale University School of Medicine, New Haven, Conn. Electronic address:

Background: Granuloma annulare (GA) is a common cutaneous inflammatory disorder characterized by macrophage accumulation and activation in skin. Its pathogenesis is poorly understood, and there are no effective treatments. The potential health implications of severe GA are unknown.

Objective: We sought to better understand GA pathogenesis and evaluate a molecularly targeted treatment approach for this disease.

Methods: We used single-cell RNA sequencing to study the immunopathogenesis of GA and also evaluated the efficacy of tofacitinib (a Janus kinase 1/3 inhibitor) in 5 patients with severe, long-standing GA in an open-label clinical trial.

Results: Using single-cell RNA sequencing, we found that in GA lesions IFN-γ production by CD4 T cells is upregulated and is associated with inflammatory polarization of macrophages and fibroblasts. In particular, macrophages upregulate oncostatin M, an IL-6 family cytokine, which appears to act on fibroblasts to alter extracellular matrix production, a hallmark of GA. IL-15 and IL-21 production appears to feed back on CD4 T cells to sustain inflammation. Treatment of 5 patients with recalcitrant GA with tofacitinib inhibited IFN-γ and oncostatin M, as well as IL-15 and IL-21, activity and resulted in clinical and histologic disease remission in 3 patients and marked improvement in the other 2. Inhibition of these effects at the molecular level paralleled the clinical improvement. Evidence of systemic inflammation is also present in some patients with severe GA and is mitigated by tofacitinib.

Conclusions: The Janus kinase-signal transducer and activator of transcription pathway is activated in GA, likely in part through the activity of IFN-γ and oncostatin M, and Janus kinase inhibitors appear to be an effective treatment.
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http://dx.doi.org/10.1016/j.jaci.2020.10.012DOI Listing
May 2021

The emerging role of Janus kinase inhibitors in the treatment of autoimmune and inflammatory diseases.

J Allergy Clin Immunol 2021 Mar 28;147(3):814-826. Epub 2020 Oct 28.

Department of Dermatology, Yale University School of Medicine, New Haven, Conn. Electronic address:

Autoimmune and inflammatory diseases are common and diverse, and they can affect nearly any organ system. Much of the pathogenesis of these diseases is related to dysregulated cytokine activity. Historically, autoimmune and inflammatory diseases have been treated with medications that nonspecifically suppress the immune system. mAbs that block the action of pathogenic cytokines emerged 2 decades ago and have become widely useful. More recently, agents that simultaneously block multiple pathogenic cytokines via inhibition of the downstream Janus kinase (JAK)-signal transducer and activator of transcription pathway have emerged and are becoming increasingly important. These small-molecule inhibitors, collectively termed JAK inhibitors, are US Food and Drug Administration-approved in a few autoimmune/inflammatory disorders and are being evaluated in many others. Here, we review the biology of the JAK-signal transducer and activator of transcription pathway and the use of JAK inhibitors to treat autoimmune and inflammatory diseases across medical subspecialties.
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http://dx.doi.org/10.1016/j.jaci.2020.10.022DOI Listing
March 2021

IL-4-BATF signaling directly modulates IL-9 producing mucosal mast cell (MMC9) function in experimental food allergy.

J Allergy Clin Immunol 2021 01 15;147(1):280-295. Epub 2020 Oct 15.

Division of Allergy and Immunology, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio; Mary H Weiser Food Allergy Center, Michigan Medicine, University of Michigan, Ann Arbor, Mich; Department of Pathology, Michigan Medicine, University of Michigan, Ann Arbor, Mich. Electronic address:

Background: This study group has previously identified IL-9-producing mucosal mast cell (MMC9) as the primary source of IL-9 to drive intestinal mastocytosis and experimental IgE-mediated food allergy. However, the molecular mechanisms that regulate the expansion of MMC9s remain unknown.

Objectives: This study hypothesized that IL-4 regulates MMC9 development and MMC9-dependent experimental IgE-mediated food allergy.

Methods: An epicutaneous sensitization model was used and bone marrow reconstitution experiments were performed to test the requirement of IL-4 receptor α (IL-4Rα) signaling on MMC9s in experimental IgE-mediated food allergy. Flow cytometric, bulk, and single-cell RNA-sequencing analyses on small intestine (SI) MMC9s were performed to illuminate MMC9 transcriptional signature and the effect of IL-4Rα signaling on MMC9 function. A bone marrow-derived MMC9 culture system was used to define IL-4-BATF signaling in MMC9 development.

Results: Epicutaneous sensitization- and bone marrow reconstitution-based models of IgE-mediated food allergy revealed an IL-4 signaling-dependent cell-intrinsic effect on SI MMC9 accumulation and food allergy severity. RNA-sequencing analysis of SI-MMC9s identified 410 gene transcripts reciprocally regulated by IL-4 signaling, including Il9 and Batf. Insilico analyses identified a 3491-gene MMC9 transcriptional signature and identified 2 transcriptionally distinct SI MMC9 populations enriched for metabolic or inflammatory programs. Employing an in vitro MMC9-culture model system showed that generation of MMC9-like cells was induced by IL-4 and this was in part dependent on BATF.

Conclusions: IL-4Rα signaling directly modulates MMC9 function and exacerbation of experimental IgE-mediated food allergic reactions. IL-4Rα regulation of MMC9s is in part BATF-dependent and occurs via modulation of metabolic transcriptional programs.
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http://dx.doi.org/10.1016/j.jaci.2020.08.043DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7856198PMC
January 2021

Macrophage scavenger receptor 1 controls Chikungunya virus infection through autophagy in mice.

Commun Biol 2020 10 8;3(1):556. Epub 2020 Oct 8.

Department of Microbiology & Immunology, School of Medicine, New York Medical College, Valhalla, NY, 10595, USA.

Macrophage scavenger receptor 1 (MSR1) mediates the endocytosis of modified low-density lipoproteins and plays an important antiviral role. However, the molecular mechanism underlying MSR1 antiviral actions remains elusive. We report that MSR1 activates autophagy to restrict infection of Chikungunya virus (CHIKV), an arthritogenic alphavirus that causes acute and chronic crippling arthralgia. Msr1 expression was rapidly upregulated after CHIKV infection in mice. Msr1 knockout mice had elevated viral loads and increased susceptibility to CHIKV arthritis along with a normal type I IFN response. Induction of LC3 lipidation by CHIKV, a marker of autophagy, was reduced in Msr1 cells. Mechanistically, MSR1 interacted with ATG12 through its cytoplasmic tail and this interaction was enhanced by CHIKV nsP1 protein. MSR1 repressed CHIKV replication through ATG5-ATG12-ATG16L1 and this was dependent on the FIP200-and-WIPI2-binding domain, but not the WD40 domain of ATG16L1. Our results elucidate an antiviral role for MSR1 involving the autophagic function of ATG5-ATG12-ATG16L1.
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http://dx.doi.org/10.1038/s42003-020-01285-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7545163PMC
October 2020

Author Correction: metabolic signaling in T cells.

Cell Res 2020 Nov;30(11):1053

Pathology and Laboratory Medicine, Children's Hospital of Philadelphia, Philadelphia, PA, USA.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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http://dx.doi.org/10.1038/s41422-020-00421-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7784931PMC
November 2020

Vibrational optical coherence tomography detects unique skin fibrotic states: Preliminary results of animal and human studies.

J Am Acad Dermatol 2020 Aug 26. Epub 2020 Aug 26.

Department of Pathology and Laboratory Medicine, Robert Wood Johnson Medical School, Rutgers, the State University of New Jersey, Piscataway, New Jersey; OptoVibronex, LLC, Mt Bethel, Pennsylvania.

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http://dx.doi.org/10.1016/j.jaad.2020.08.091DOI Listing
August 2020

A special collection of reviews on frontiers in immunology.

Cell Res 2020 10;30(10):827-828

Department of Immunobiology, Yale School of Medicine, New Haven, CT, USA.

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http://dx.doi.org/10.1038/s41422-020-00403-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7454546PMC
October 2020

Sulfamethoxazole drug stress upregulates antioxidant immunomodulatory metabolites in Escherichia coli.

Nat Microbiol 2020 11 27;5(11):1319-1329. Epub 2020 Jul 27.

Department of Chemistry, Yale University, New Haven, CT, USA.

Escherichia coli is an important model organism in microbiology and a prominent member of the human microbiota. Environmental isolates readily colonize the gastrointestinal tract of humans and other animals, and they can serve diverse probiotic, commensal and pathogenic roles in the host. Although certain strains have been associated with the severity of inflammatory bowel disease (IBD), the diverse immunomodulatory phenotypes remain largely unknown at the molecular level. Here, we decode a previously unknown E. coli metabolic pathway that produces a family of hybrid pterin-phenylpyruvate conjugates, which we named the colipterins. The metabolites are upregulated by subinhibitory levels of the antifolate sulfamethoxazole, which is used to treat infections including in patients with IBD. The genes folX/M and aspC/tyrB involved in monapterin biosynthesis and aromatic amino acid transamination, respectively, were required to initiate the colipterin pathway. We show that the colipterins are antioxidants, harbour diverse immunological activities in primary human tissues, activate anti-inflammatory interleukin-10 and improve colitis symptoms in a colitis mouse model. Our study defines an antifolate stress response in E. coli and links its associated metabolites to a major immunological marker of IBD.
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http://dx.doi.org/10.1038/s41564-020-0763-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7581551PMC
November 2020

Metabolic signaling in T cells.

Cell Res 2020 08 24;30(8):649-659. Epub 2020 Jul 24.

Pathology and Laboratory Medicine, Children's Hospital of Philadelphia, Philadelphia, PA, USA.

The maintenance of organismal homeostasis requires partitioning and transport of biochemical molecules between organ systems, their composite cells, and subcellular organelles. Although transcriptional programming undeniably defines the functional state of cells and tissues, underlying biochemical networks are intricately intertwined with transcriptional, translational, and post-translational regulation. Studies of the metabolic regulation of immunity have elegantly illustrated this phenomenon. The cells of the immune system interface with a diverse set of environmental conditions. Circulating immune cells perfuse peripheral organs in the blood and lymph, patrolling for pathogen invasion. Resident immune cells remain in tissues and play more newly appreciated roles in tissue homeostasis and immunity. Each of these cell populations interacts with unique and dynamic tissue environments, which vary greatly in biochemical composition. Furthermore, the effector response of immune cells to a diverse set of activating cues requires unique cellular adaptations to supply the requisite biochemical landscape. In this review, we examine the role of spatial partitioning of metabolic processes in immune function. We focus on studies of lymphocyte metabolism, with reference to the greater immunometabolism literature when appropriate to illustrate this concept.
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http://dx.doi.org/10.1038/s41422-020-0379-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7395146PMC
August 2020

CFTR is a negative regulator of γδ T cell IFN-γ production and antitumor immunity.

Cell Mol Immunol 2020 Jul 15. Epub 2020 Jul 15.

Zhuhai Precision Medical Center, Zhuhai People's Hospital (Zhuhai Hospital Affiliated with Jinan University), Jinan University, Zhuhai, 519000, Guangdong, China.

CFTR, a chloride channel and ion channel regulator studied mostly in epithelial cells, has been reported to participate in immune regulation and likely affect the risk of cancer development. However, little is known about the effects of CFTR on the differentiation and function of γδ T cells. In this study, we observed that CFTR was functionally expressed on the cell surface of γδ T cells. Genetic deletion and pharmacological inhibition of CFTR both increased IFN-γ release by peripheral γδ T cells and potentiated the cytolytic activity of these cells against tumor cells both in vitro and in vivo. Interestingly, the molecular mechanisms underlying the regulation of γδ T cell IFN-γ production by CFTR were either TCR dependent or related to Ca influx. CFTR was recruited to TCR immunological synapses and attenuated Lck-P38 MAPK-c-Jun signaling. In addition, CFTR was found to modulate TCR-induced Ca influx and membrane potential (V)-induced Ca influx and subsequently regulate the calcineurin-NFATc1 signaling pathway in γδ T cells. Thus, CFTR serves as a negative regulator of IFN-γ production in γδ T cells and the function of these cells in antitumor immunity. Our investigation suggests that modification of the CFTR activity of γδ T cells may be a potential immunotherapeutic strategy for cancer.
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http://dx.doi.org/10.1038/s41423-020-0499-3DOI Listing
July 2020

The induction and function of the anti-inflammatory fate of T17 cells.

Nat Commun 2020 07 3;11(1):3334. Epub 2020 Jul 3.

Department of General, Visceral and Thoracic Surgery, University Medical Center Hamburg-Eppendorf, 20246, Hamburg, Germany.

T17 cells exemplify environmental immune adaptation: they can acquire both a pathogenic and an anti-inflammatory fate. However, it is not known whether the anti-inflammatory fate is merely a vestigial trait, or whether it serves to preserve the integrity of the host tissues. Here we show that the capacity of T17 cells to acquire an anti-inflammatory fate is necessary to sustain immunological tolerance, yet it impairs immune protection against S. aureus. Additionally, we find that TGF-β signalling via Smad3/Smad4 is sufficient for the expression of the anti-inflammatory cytokine, IL-10, in T17 cells. Our data thus indicate a key function of T17 cell plasticity in maintaining immune homeostasis, and dissect the molecular mechanisms explaining the functional flexibility of T17 cells with regard to environmental changes.
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http://dx.doi.org/10.1038/s41467-020-17097-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7335205PMC
July 2020

IL22BP Mediates the Antitumor Effects of Lymphotoxin Against Colorectal Tumors in Mice and Humans.

Gastroenterology 2020 10 22;159(4):1417-1430.e3. Epub 2020 Jun 22.

Georgia Cancer Center, Augusta University, Augusta, Georgia.

Background & Aims: Unregulated activity of interleukin (IL) 22 promotes intestinal tumorigenesis in mice. IL22 binds the antagonist IL22 subunit alpha 2 (IL22RA2, also called IL22BP). We studied whether alterations in IL22BP contribute to colorectal carcinogenesis in humans and mice.

Methods: We obtained tumor and nontumor tissues from patients with colorectal cancer (CRC) and measured levels of cytokines by quantitative polymerase chain reaction, flow cytometry, and immunohistochemistry. We measured levels of Il22bp messenger RNA in colon tissues from wild-type, Tnf, Lta, and Ltb mice. Mice were given azoxymethane and dextran sodium sulfate to induce colitis and associated cancer or intracecal injections of MC38 tumor cells. Some mice were given inhibitors of lymphotoxin beta receptor (LTBR). Intestine tissues were analyzed by single-cell sequencing to identify cell sources of lymphotoxin. We performed immunohistochemistry analysis of colon tissue microarrays from patients with CRC (1475 tissue cores, contained tumor and nontumor tissues) and correlated levels of IL22BP with patient survival times.

Results: Levels of IL22BP were decreased in human colorectal tumors, compared with nontumor tissues, and correlated with levels of lymphotoxin. LTBR signaling was required for expression of IL22BP in colon tissues of mice. Wild-type mice given LTBR inhibitors had an increased tumor burden in both models, but LTBR inhibitors did not increase tumor growth in Il22bp mice. Lymphotoxin directly induced expression of IL22BP in cultured human monocyte-derived dendritic cells via activation of nuclear factor κB. Reduced levels of IL22BP in colorectal tumor tissues were associated with shorter survival times of patients with CRC.

Conclusions: Lymphotoxin signaling regulates expression of IL22BP in colon; levels of IL22BP are reduced in human colorectal tumors, associated with shorter survival times. LTBR signaling regulates expression of IL22BP in colon tumors in mice and cultured human dendritic cells. Patients with colorectal tumors that express low levels of IL22BP might benefit from treatment with an IL22 antagonist.
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http://dx.doi.org/10.1053/j.gastro.2020.06.033DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7607422PMC
October 2020