Publications by authors named "Katherine A Fitzgerald"

250 Publications

cGAS-STING Pathway Does Not Promote Autoimmunity in Murine Models of SLE.

Front Immunol 2021 29;12:605930. Epub 2021 Mar 29.

Program in Innate Immunity, Division of Infectious Diseases and Immunology, Department of Medicine, University of Massachusetts Medical School, Worcester, MA, United States.

Detection of DNA is an important determinant of host-defense but also a driver of autoinflammatory and autoimmune diseases. Failure to degrade self-DNA in DNAseII or III(TREX1)-deficient mice results in activation of the cGAS-STING pathway. Deficiency of cGAS or STING in these models ameliorates disease manifestations. However, the contribution of the cGAS-STING pathway, relative to endosomal TLRs, in systemic lupus erythematosus (SLE) is controversial. In fact, STING deficiency failed to rescue, and actually exacerbated, disease manifestations in Fas-deficient SLE-prone mice. We have now extended these observations to a chronic model of SLE induced by the i.p. injection of TMPD (pristane). We found that both cGAS- and STING-deficiency not only failed to rescue mice from TMPD-induced SLE, but resulted in increased autoantibody production and higher proteinuria levels compared to cGAS STING sufficient mice. Further, we generated cGASFas mice on a pure MRL/Fas background using Crispr/Cas9 and found slightly exacerbated, and not attenuated, disease. We hypothesized that the cGAS-STING pathway constrains TLR activation, and thereby limits autoimmune manifestations in these two models. Consistent with this premise, mice lacking cGAS and Unc93B1 or STING and Unc93B1 developed minimal systemic autoimmunity as compared to cGAS or STING single knock out animals. Nevertheless, TMPD-driven lupus in B6 mice was abrogated upon AAV-delivery of DNAse I, implicating a DNA trigger. Overall, this study demonstrated that the cGAS-STING pathway does not promote systemic autoimmunity in murine models of SLE. These data have important implications for cGAS-STING-directed therapies being developed for the treatment of systemic autoimmunity.
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http://dx.doi.org/10.3389/fimmu.2021.605930DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8040952PMC
March 2021

Lung Epithelial Cell Transcriptional Regulation as a Factor in COVID-19 Associated Coagulopathies.

Am J Respir Cell Mol Biol 2021 Mar 19. Epub 2021 Mar 19.

Brown University Division of Biology and Medicine, 174279, MMI, Providence, Rhode Island, United States;

SARS-CoV-2 has rapidly become a global pandemic. In addition to the acute pulmonary symptoms of COVID-19 (the disease associated with SARS-CoV-2 infection), pulmonary and distal coagulopathies have caused morbidity and mortality in many patients. Currently, the molecular pathogenesis underlying COVID-19 associated coagulopathies are unknown. Identifying the molecular basis of how SARS-CoV-2 drives coagulation is essential to mitigating short and long term thrombotic risks of sick and recovered COVID-19 patients. We aimed to perform coagulation focused transcriptome analysis of in vitro infected primary respiratory epithelial cells, patient derived bronchial alveolar lavage (BALF) cells, and circulating immune cells during SARS-CoV-2 infection. Our objective was to identify transcription mediated signaling networks driving coagulopathies associated with COVID-19. We analyzed recently published experimentally and clinically derived bulk or single cell RNA sequencing datasets of SARS-CoV-2 infection to identify changes in transcriptional regulation of blood coagulation. We also confirmed that the transcriptional expression of a key coagulation regulator was recapitulated at the protein level. We specifically focused our analysis on lung tissue expressed genes regulating the extrinsic coagulation cascade and the plasminogen activation system. Analyzing transcriptomic data of in vitro infected normal human bronchial epithelial (NHBE) cells and patient derived BALF samples revealed that SARS-CoV-2 infection induces the extrinsic blood coagulation cascade and suppresses the plasminogen activation system. We also performed in vitro SARS-CoV-2 infection experiments on primary human lung epithelial cells to confirm that transcriptional upregulation of Tissue Factor, the extrinsic coagulation cascade master regulator, manifested at the protein level. Further, infection of NHBEs with influenza A virus (IAV) did not drive key regulators of blood coagulation in a similar manner as SARS-CoV-2. Additionally, peripheral blood mononuclear cells (PBMCs) did not differentially express genes regulating the extrinsic coagulation cascade or plasminogen activation system during SARS-CoV-2 infection, suggesting that they are not directly inducing coagulopathy through these pathways. The hyper-activation of the extrinsic blood coagulation cascade and the suppression of the plasminogen activation system in SARS-CoV-2 infected epithelial cells may drive diverse coagulopathies in the lung and distal organ systems. Understanding how hosts drive such transcriptional changes with SARS-CoV-2 infection may enable the design of host-directed therapeutic strategies to treat COVID-19 and other coronaviruses inducing hyper-coagulation. This article is open access and distributed under the terms of the Creative Commons Attribution Non-Commercial No Derivatives License 4.0 (http://creativecommons.org/licenses/by-nc-nd/4.0/).
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http://dx.doi.org/10.1165/rcmb.2020-0453OCDOI Listing
March 2021

Overcoming innate immune barriers that impede AAV gene therapy vectors.

J Clin Invest 2021 Jan;131(1)

Horae Gene Therapy Center.

The field of gene therapy has made considerable progress over the past several years. Adeno-associated virus (AAV) vectors have emerged as promising and attractive tools for in vivo gene therapy. Despite the recent clinical successes achieved with recombinant AAVs (rAAVs) for therapeutics, host immune responses against the vector and transgene product have been observed in numerous preclinical and clinical studies. These outcomes have hampered the advancement of AAV gene therapies, preventing them from becoming fully viable and safe medicines. The human immune system is multidimensional and complex. Both the innate and adaptive arms of the immune system seem to play a concerted role in the response against rAAVs. While most efforts have been focused on the role of adaptive immunity and developing ways to overcome it, the innate immune system has also been found to have a critical function. Innate immunity not only mediates the initial response to the vector, but also primes the adaptive immune system to launch a more deleterious attack against the foreign vector. This Review highlights what is known about innate immune responses against rAAVs and discusses potential strategies to circumvent these pathways.
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http://dx.doi.org/10.1172/JCI143780DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7773343PMC
January 2021

The long non-coding RNA LUCAT1 is a negative feedback regulator of interferon responses in humans.

Nat Commun 2020 12 11;11(1):6348. Epub 2020 Dec 11.

Program in Innate Immunity, University of Massachusetts Medical School, Worcester, MA, 01605, USA.

Long non-coding RNAs are important regulators of biological processes including immune responses. The immunoregulatory functions of lncRNAs have been revealed primarily in murine models with limited understanding of lncRNAs in human immune responses. Here, we identify lncRNA LUCAT1 which is upregulated in human myeloid cells stimulated with lipopolysaccharide and other innate immune stimuli. Targeted deletion of LUCAT1 in myeloid cells increases expression of type I interferon stimulated genes in response to LPS. By contrast, increased LUCAT1 expression results in a reduction of the inducible ISG response. In activated cells, LUCAT1 is enriched in the nucleus where it associates with chromatin. Further, LUCAT1 limits transcription of interferon stimulated genes by interacting with STAT1 in the nucleus. Together, our study highlights the role of the lncRNA LUCAT1 as a post-induction feedback regulator which functions to restrain the immune response in human cells.
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http://dx.doi.org/10.1038/s41467-020-20165-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7733444PMC
December 2020

Loosening the grip on nuclear cGAS.

Nat Genet 2020 12;52(12):1269-1270

Program in Innate Immunity, Division of Infectious Diseases and Immunology, Department of Medicine, University of Massachusetts Medical School, Worcester, MA, USA.

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http://dx.doi.org/10.1038/s41588-020-00746-2DOI Listing
December 2020

Ensuring vaccine safety.

Science 2020 12 17;370(6522):1274-1275. Epub 2020 Nov 17.

Department of Microbiology and Immunology, National Emerging Infectious Diseases Laboratories, Boston University School of Medicine, Boston, MA, USA.

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http://dx.doi.org/10.1126/science.abf0357DOI Listing
December 2020

Author Correction: Caspase-8 mediates inflammation and disease in rodent malaria.

Nat Commun 2020 Nov 3;11(1):5673. Epub 2020 Nov 3.

Instituto Rene Rachou, FIOCRUZ-MG, Belo Horizonte, MG, 30190-002, Brazil.

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/s41467-020-19620-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7641112PMC
November 2020

HDAC6 mediates an aggresome-like mechanism for NLRP3 and pyrin inflammasome activation.

Science 2020 09;369(6510)

Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA.

Inflammasomes are supramolecular complexes that play key roles in immune surveillance. This is accomplished by the activation of inflammatory caspases, which leads to the proteolytic maturation of interleukin 1β (IL-1β) and pyroptosis. Here, we show that nucleotide-binding domain, leucine-rich repeat, and pyrin domain-containing protein 3 (NLRP3)- and pyrin-mediated inflammasome assembly, caspase activation, and IL-1β conversion occur at the microtubule-organizing center (MTOC). Furthermore, the dynein adapter histone deacetylase 6 (HDAC6) is indispensable for the microtubule transport and assembly of these inflammasomes both in vitro and in mice. Because HDAC6 can transport ubiquitinated pathological aggregates to the MTOC for aggresome formation and autophagosomal degradation, its role in NLRP3 and pyrin inflammasome activation also provides an inherent mechanism for the down-regulation of these inflammasomes by autophagy. This work suggests an unexpected parallel between the formation of physiological and pathological aggregates.
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http://dx.doi.org/10.1126/science.aas8995DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7814939PMC
September 2020

Caspase-8 mediates inflammation and disease in rodent malaria.

Nat Commun 2020 09 14;11(1):4596. Epub 2020 Sep 14.

Instituto Rene Rachou, FIOCRUZ-MG, Belo Horizonte, MG, 30190-002, Brazil.

Earlier studies indicate that either the canonical or non-canonical pathways of inflammasome activation have a limited role on malaria pathogenesis. Here, we report that caspase-8 is a central mediator of systemic inflammation, septic shock in the Plasmodium chabaudi-infected mice and the P. berghei-induced experimental cerebral malaria (ECM). Importantly, our results indicate that the combined deficiencies of caspases-8/1/11 or caspase-8/gasdermin-D (GSDM-D) renders mice impaired to produce both TNFα and IL-1β and highly resistant to lethality in these models, disclosing a complementary, but independent role of caspase-8 and caspases-1/11/GSDM-D in the pathogenesis of malaria. Further, we find that monocytes from malaria patients express active caspases-1, -4 and -8 suggesting that these inflammatory caspases may also play a role in the pathogenesis of human disease.
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http://dx.doi.org/10.1038/s41467-020-18295-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7490701PMC
September 2020

Succination inactivates gasdermin D and blocks pyroptosis.

Science 2020 09 20;369(6511):1633-1637. Epub 2020 Aug 20.

Program in Innate Immunity, Department of Medicine, University of Massachusetts Medical School, Worcester, MA 01605, USA.

Activated macrophages undergo a metabolic switch to aerobic glycolysis, accumulating Krebs' cycle intermediates that alter transcription of immune response genes. We extended these observations by defining fumarate as an inhibitor of pyroptotic cell death. We found that dimethyl fumarate (DMF) delivered to cells or endogenous fumarate reacts with gasdermin D (GSDMD) at critical cysteine residues to form S-(2-succinyl)-cysteine. GSDMD succination prevents its interaction with caspases, limiting its processing, oligomerization, and capacity to induce cell death. In mice, the administration of DMF protects against lipopolysaccharide shock and alleviates familial Mediterranean fever and experimental autoimmune encephalitis by targeting GSDMD. Collectively, these findings identify GSDMD as a target of fumarate and reveal a mechanism of action for fumarate-based therapeutics that include DMF, for the treatment of multiple sclerosis.
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http://dx.doi.org/10.1126/science.abb9818DOI Listing
September 2020

Long non-coding RNAs in antiviral immunity.

Semin Cell Dev Biol 2021 Mar 21;111:126-134. Epub 2020 Jun 21.

Program in Innate Immunity, Department of Medicine, University of Massachusetts Medical School, Worcester, MA, United States. Electronic address:

Tight regulation of the immune response is fundamental for efficient pathogen clearance and to prevent excessive inflammation. Long non-coding RNAs (lncRNAs) have emerged as potent regulators of the innate and adaptive immune responses to viral pathogens. Host-derived lncRNAs control the differentiation and polarization of immune cell populations and the production of cytokines, interferons and antiviral factors. This review provides an updated overview of lncRNAs that modulate viral replication or pathogenesis. Beyond that, viruses have developed lncRNA-based strategies to mask themselves from immune detection and evade antiviral immunity. A deeper understanding of lncRNA biology in the context of host-pathogen interactions may unveil new treatment strategies in the near future.
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http://dx.doi.org/10.1016/j.semcdb.2020.06.009DOI Listing
March 2021

Inflammasomes.

Curr Biol 2020 06;30(12):R689-R694

Program in Innate Immunity, Division of Infectious Diseases and Immunology, Department of Medicine, University of Massachusetts Medical School, Worcester, MA 01605, USA. Electronic address:

Ketelut-Carneiro and Fitzgerald discuss the basic concepts about inflammasome composition, assembly and activation, effector functions and role in infection and inflammatory disease.
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http://dx.doi.org/10.1016/j.cub.2020.04.065DOI Listing
June 2020

Toll-like Receptors and the Control of Immunity.

Cell 2020 03 11;180(6):1044-1066. Epub 2020 Mar 11.

Harvard Medical School and Division of Gastroenterology, Boston Children's Hospital, Boston, MA, USA. Electronic address:

The study of innate immunity and its link to inflammation and host defense encompasses diverse areas of biology, ranging from genetics and biophysics to signal transduction and physiology. Central to our understanding of these events are the Toll-like receptors (TLRs), an evolutionarily ancient family of pattern recognition receptors. Herein, we describe the mechanisms and consequences of TLR-mediated signal transduction with a focus on themes identified in the TLR pathways that also explain the operation of other immune signaling pathways. These themes include the detection of conserved microbial structures to identify infectious agents and the use of supramolecular organizing centers (SMOCs) as signaling organelles that ensure digital cellular responses. Further themes include mechanisms of inducible gene expression, the coordination of gene regulation and metabolism, and the influence of these activities on adaptive immunity. Studies in these areas have informed the development of next-generation therapeutics, thus ensuring a bright future for research in this area.
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http://dx.doi.org/10.1016/j.cell.2020.02.041DOI Listing
March 2020

Direct Binding to NLRP3 Pyrin Domain as a Novel Strategy to Prevent NLRP3-Driven Inflammation and Gouty Arthritis.

Arthritis Rheumatol 2020 07 27;72(7):1192-1202. Epub 2020 May 27.

College of Pharmacy, The Catholic University of Korea, Bucheon, Republic of Korea.

Objective: The NLRP3 inflammasome is closely linked to the pathophysiology of a wide range of inflammatory diseases. This study was undertaken to identify small molecules that directly bind to NLRP3 in order to develop pharmacologic interventions for NLRP3-related diseases.

Methods: A structure-based virtual screening analysis was performed with ~62,800 compounds to select efficient NLRP3 inhibitors. The production of caspase 1-p10 and interleukin-1β (IL-1β) was measured by immunoblotting and enzyme-linked immunosorbent assay to examine NLRP3 inflammasome activation. Two gouty arthritis models and an air pouch inflammation model induced by monosodium urate monohydrate (MSU) crystal injection were used for in vivo experiments. Primary synovial fluid cells from gout patients were used to determine the relevance of NLRP3 inflammasome inhibition in human gout.

Results: Beta-carotene (provitamin A) suppressed the NLRP3 inflammasome activation induced by various activators, including MSU crystals, in mouse bone marrow-derived primary macrophages (P < 0.05). Surface plasmon resonance analysis demonstrated the direct binding of β-carotene to the pyrin domain (PYD) of NLRP3 (K = 3.41 × 10 ). Molecular modeling and mutation assays revealed the interaction mode between β-carotene and the NLRP3 PYD. Inflammatory symptoms induced by MSU crystals were attenuated by oral administration of β-carotene in gouty arthritis mouse models (P < 0.05), correlating with its suppressive effects on the NLRP3 inflammasome in inflamed tissues. Furthermore, β-carotene reduced IL-1β secretion from human synovial fluid cells isolated from gout patients (P < 0.05), showing its inhibitory efficacy in human gout.

Conclusion: Our results present β-carotene as a selective and direct inhibitor of NLRP3, and the binding of β-carotene to NLRP3 PYD as a novel pharmacologic strategy to combat NLRP3 inflammasome-driven diseases, including gouty arthritis.
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http://dx.doi.org/10.1002/art.41245DOI Listing
July 2020

Long Non-coding RNA LincRNA-EPS Inhibits Host Defense Against Infection.

Front Cell Infect Microbiol 2019 22;9:481. Epub 2020 Jan 22.

Department of Immunology, UConn Health School of Medicine, Farmington, CT, United States.

Long non-coding RNAs (lncRNAs) have emerged as key regulators of gene expression in several biological systems. The long intergenic RNA-erythroid pro-survival (lincRNA-EPS) has been shown to play a critical role in restraining inflammatory gene expression. However, the function of lincRNA-EPS during bacterial infections remains unknown. Here, we demonstrate that following infection with the intracellular bacterium , both mouse macrophages and dendritic cells lacking lincRNA-EPS exhibit an enhanced expression of proinflammatory cytokine genes, as well as an increased expression of the inducible nitric oxide synthase ( and nitric oxide (NO) production. Importantly, we found that lincRNA-EPS mice intraperitoneally infected with exhibit lower bacterial burdens in spleen and liver and produce more NO than control mice. Furthermore, lincRNA-EPS mice are less susceptible to a lethal dose of than wild type (WT) mice. Collectively these findings show that lincRNA-EPS suppresses host protective NO expression and impairs the host defense against infection.
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http://dx.doi.org/10.3389/fcimb.2019.00481DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6987077PMC
August 2020

TLR2 Dimerization Blockade Allows Generation of Homeostatic Intestinal Macrophages under Acute Colitis Challenge.

J Immunol 2020 02 27;204(3):707-717. Epub 2019 Dec 27.

Department of Immunology, Weizmann Institute of Science, Rehovot 76100, Israel;

Recruited blood monocytes contribute to the establishment, perpetuation, and resolution of tissue inflammation. Specifically, in the inflamed intestine, monocyte ablation was shown to ameliorate colitis scores in preclinical animal models. However, the majority of intestinal macrophages that seed the healthy gut are also monocyte derived. Monocyte ablation aimed to curb inflammation would therefore likely interfere with intestinal homeostasis. In this study, we used a TLR2 -membrane peptide that blocks TLR2 dimerization that is critical for TLR2/1 and TLR2/6 heterodimer signaling to blunt inflammation in a murine colitis model. We show that although the TLR2 peptide treatment ameliorated colitis, it allowed recruited monocytes to give rise to macrophages that lack the detrimental proinflammatory gene signature and reduced potentially damaging neutrophil infiltrates. Finally, we demonstrate TLR blocking activity of the peptide on in vitro cultured human monocyte-derived macrophages. Collectively, we provide a significantly improved anti-inflammatory TLR2 peptide and critical insights in its mechanism of action toward future potential use in the clinic.
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http://dx.doi.org/10.4049/jimmunol.1900470DOI Listing
February 2020

A Mitochondrial Micropeptide Is Required for Activation of the Nlrp3 Inflammasome.

J Immunol 2020 01 13;204(2):428-437. Epub 2019 Dec 13.

Program in Innate Immunity, Department of Medicine, University of Massachusetts Medical School, Worcester, MA 01605; and

Functional peptides encoded by short open reading frames are emerging as important mediators of fundamental biological processes. In this study, we identified a micropeptide produced from a putative long noncoding RNA (lncRNAs) that is important in controlling innate immunity. By studying lncRNAs in mice macrophages, we identified lncRNA 1810058I24Rik, which was downregulated in both human and murine myeloid cells exposed to LPS as well as other TLR ligands and inflammatory cytokines. Analysis of lncRNA 1810058I24Rik subcellular localization revealed that this transcript was localized in the cytosol, prompting us to evaluate its coding potential. In vitro translation with S-labeled methionine resulted in translation of a 47 aa micropeptide. Microscopy and subcellular fractionation studies in macrophages demonstrated endogenous expression of this peptide on the mitochondrion. We thus named this gene mitochondrial micropeptide-47 (Mm47). Crispr-Cas9-mediated deletion of , as well as small interfering RNA studies in mice primary macrophages, showed that the transcriptional response downstream of TLR4 was intact in cells lacking Mm47. In contrast, -deficient or knockdown cells were compromised for Nlrp3 inflammasome responses. Activation of Nlrc4 or Aim2 inflammasomes were intact in cells lacking Mm47. This study therefore identifies, to our knowledge, a novel mitochondrial micropeptide Mm47 that is required for the activation of the Nlrp3 inflammasome. This work further highlights the functional activity of short open reading frame-encoded peptides and underscores their importance in innate immunity.
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http://dx.doi.org/10.4049/jimmunol.1900791DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7370245PMC
January 2020

hnRNPA2B1: Fueling Antiviral Immunity from the Nucleus.

Mol Cell 2019 10;76(1):8-10

University of Massachusetts Medical School, Worcester, MA 01655, USA. Electronic address:

In a recently published article in Science, Cao and colleagues (Wang et al., 2019) identify hnRNPA2B1 as a new DNA-binding protein that initiates and amplifies antiviral immunity, unveiling a new facet of DNA recognition in the nucleus.
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http://dx.doi.org/10.1016/j.molcel.2019.09.021DOI Listing
October 2019

Gasdermins and their role in immunity and inflammation.

J Exp Med 2019 11 23;216(11):2453-2465. Epub 2019 Sep 23.

Program in Innate Immunity, Division of Infectious Diseases and Immunology, Department of Medicine, University of Massachusetts Medical School, Worcester, MA

The gasdermins are a family of pore-forming proteins recently implicated in the immune response. One of these proteins, gasdermin D (GSDMD), has been identified as the executioner of pyroptosis, an inflammatory form of lytic cell death that is induced upon formation of caspase-1-activating inflammasomes. The related proteins GSDME and GSDMA have also been implicated in autoimmune diseases and certain cancers. Most gasdermin proteins are believed to have pore-forming capabilities. The best-studied member, GSDMD, controls the release of the proinflammatory cytokines IL-1ß and IL-18 and pyroptotic cell death. Because of its potential as a driver of inflammation in septic shock and autoimmune diseases, GSDMD represents an attractive drug target. In this review, we discuss the gasdermin proteins with particular emphasis on GSDMD and its mechanism of action and biological significance.
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http://dx.doi.org/10.1084/jem.20190545DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6829603PMC
November 2019

Assembling the Inflammasome, Piece by Piece.

J Immunol 2019 09;203(5):1093-1094

Program in Innate Immunity, Division of Infectious Diseases and Immunology, Department of Medicine, University of Massachusetts Medical School, Worcester, MA 01605

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http://dx.doi.org/10.4049/jimmunol.1900764DOI Listing
September 2019

DNA sensing by the cGAS-STING pathway in health and disease.

Nat Rev Genet 2019 11 29;20(11):657-674. Epub 2019 Jul 29.

Program in Innate Immunity, Division of Infectious Diseases and Immunology, Department of Medicine, University of Massachusetts Medical School, Worcester, MA, USA.

The detection of pathogens through nucleic acid sensors is a defining principle of innate immunity. RNA-sensing and DNA-sensing receptors sample subcellular compartments for foreign nucleic acids and, upon recognition, trigger immune signalling pathways for host defence. Over the past decade, our understanding of how the recognition of nucleic acids is coupled to immune gene expression has advanced considerably, particularly for the DNA-sensing receptor cyclic GMP-AMP synthase (cGAS) and its downstream signalling effector stimulator of interferon genes (STING), as well as the molecular components and regulation of this pathway. Moreover, the ability of self-DNA to engage cGAS has emerged as an important mechanism fuelling the development of inflammation and implicating the cGAS-STING pathway in human inflammatory diseases and cancer. This detailed mechanistic and biological understanding is paving the way for the development and clinical application of pharmacological agonists and antagonists in the treatment of chronic inflammation and cancer.
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http://dx.doi.org/10.1038/s41576-019-0151-1DOI Listing
November 2019

Control of antiviral innate immune response by protein geranylgeranylation.

Sci Adv 2019 05 29;5(5):eaav7999. Epub 2019 May 29.

Division of Rheumatology and Immunology, Department of Medicine, Duke University School of Medicine, 207 Research Drive, Durham, NC 27710, USA.

The mitochondrial antiviral signaling protein (MAVS) orchestrates host antiviral innate immune response to RNA virus infection. However, how MAVS signaling is controlled to eradicate virus while preventing self-destructive inflammation remains obscure. Here, we show that protein geranylgeranylation, a posttranslational lipid modification of proteins, limits MAVS-mediated immune signaling by targeting Rho family small guanosine triphosphatase Rac1 into the mitochondria-associated endoplasmic reticulum (ER) membranes (MAMs) at the mitochondria-ER junction. Protein geranylgeranylation and subsequent palmitoylation promote Rac1 translocation into MAMs upon viral infection. MAM-localized Rac1 limits MAVS' interaction with E3 ligase Trim31 and hence inhibits MAVS ubiquitination, aggregation, and activation. Rac1 also facilitates the recruitment of caspase-8 and cFLIP to the MAVS signalosome and the subsequent cleavage of Ripk1 that terminates MAVS signaling. Consistently, mice with myeloid deficiency of protein geranylgeranylation showed improved survival upon influenza A virus infection. Our work revealed a critical role of protein geranylgeranylation in regulating antiviral innate immune response.
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http://dx.doi.org/10.1126/sciadv.aav7999DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6541464PMC
May 2019

HiChIRP reveals RNA-associated chromosome conformation.

Nat Methods 2019 06 27;16(6):489-492. Epub 2019 May 27.

Center for Personal Dynamic Regulomes, Stanford University School of Medicine, Stanford, CA, USA.

Modular domains of long non-coding RNAs can serve as scaffolds to bring distant regions of the linear genome into spatial proximity. Here, we present HiChIRP, a method leveraging bio-orthogonal chemistry and optimized chromosome conformation capture conditions, which enables interrogation of chromatin architecture focused around a specific RNA of interest down to approximately ten copies per cell. HiChIRP of three nuclear RNAs reveals insights into promoter interactions (7SK), telomere biology (telomerase RNA component) and inflammatory gene regulation (lincRNA-EPS).
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http://dx.doi.org/10.1038/s41592-019-0407-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6638558PMC
June 2019

Cell Survival and Cytokine Release after Inflammasome Activation Is Regulated by the Toll-IL-1R Protein SARM.

Immunity 2019 06 7;50(6):1412-1424.e6. Epub 2019 May 7.

School of Biochemistry and Immunology, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin 2, Ireland. Electronic address:

Assembly of inflammasomes after infection or injury leads to the release of interleukin-1β (IL-1β) and to pyroptosis. After inflammasome activation, cells either pyroptose or enter a hyperactivated state defined by IL-1β secretion without cell death, but what controls these different outcomes is unknown. Here, we show that removal of the Toll-IL-1R protein SARM from macrophages uncouples inflammasome-dependent cytokine release and pyroptosis, whereby cells displayed increased IL-1β production but reduced pyroptosis. Correspondingly, increasing SARM in cells caused less IL-1β release and more pyroptosis. SARM suppressed IL-1β by directly restraining the NLRP3 inflammasome and, hence, caspase-1 activation. Consistent with a role for SARM in pyroptosis, Sarm1 mice were protected from lipopolysaccharide (LPS)-stimulated sepsis. Pyroptosis-inducing, but not hyperactivating, NLRP3 stimulants caused SARM-dependent mitochondrial depolarization. Thus, SARM-dependent mitochondrial depolarization distinguishes NLRP3 activators that cause pyroptosis from those that do not, and SARM modulation represents a cell-intrinsic mechanism to regulate cell fate after inflammasome activation.
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http://dx.doi.org/10.1016/j.immuni.2019.04.005DOI Listing
June 2019

Hierarchy of clinical manifestations in SAVI N153S and V154M mouse models.

Proc Natl Acad Sci U S A 2019 04 3;116(16):7941-7950. Epub 2019 Apr 3.

Program in Innate Immunity, Division of Infectious Diseases and Immunology, Department of Medicine, University of Massachusetts Medical School, Worcester, MA 01605;

Studies over the past decade have revealed a central role for innate immune sensors in autoimmune and autoinflammatory diseases. cGAS, a cytosolic DNA sensor, detects both foreign and host DNA and generates a second-messenger cGAMP, which in turn binds and activates stimulator of IFN genes (STING), leading to induction of type I interferons and inflammatory cytokines. Recently, gain-of-function mutations in STING have been identified in patients with STING-associated vasculopathy with onset in infancy (SAVI). SAVI patients present with early-onset systemic inflammation and interstitial lung disease, resulting in pulmonary fibrosis and respiratory failure. Here, we describe two independent SAVI mouse models, harboring the two most common mutations found in patients. A direct comparison of these strains reveals a hierarchy of immune abnormalities, lung inflammation and fibrosis, which do not depend on either IFN-α/β receptor signaling or mixed lineage kinase domain-like pseudokinase (MLKL)-dependent necroptotic cell death pathways. Furthermore, radiation chimera experiments reveal how bone marrow from the V154M mutant mice transfer disease to the WT host, whereas the N153S does not, indicating mutation-specific disease outcomes. Moreover, using radiation chimeras we find that T cell lymphopenia depends on T cell-intrinsic expression of the SAVI mutation. Collectively, these mutant mice recapitulate many of the disease features seen in SAVI patients and highlight mutation-specific functions of STING that shed light on the heterogeneity observed in SAVI patients.
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http://dx.doi.org/10.1073/pnas.1818281116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6475399PMC
April 2019

CNBP controls IL-12 gene transcription and Th1 immunity.

J Exp Med 2018 12 15;215(12):3136-3150. Epub 2018 Nov 15.

Program in Innate Immunity, Department of Medicine, University of Massachusetts Medical School, Worcester, MA

An inducible program of inflammatory gene expression is a hallmark of antimicrobial defenses. Recently, cellular nucleic acid-binding protein (CNBP) was identified as a regulator of nuclear factor-kappaB (NF-κB)-dependent proinflammatory cytokine gene expression. Here, we generated mice lacking CNBP and found that CNBP regulates a very restricted gene signature that includes IL-12β. CNBP resides in the cytosol of macrophages and translocates to the nucleus in response to diverse microbial pathogens and pathogen-derived products. -deficient macrophages induced canonical NF-κB/Rel signaling normally but were impaired in their ability to control the activation of c-Rel, a key driver of IL-12β gene transcription. The nuclear translocation and DNA-binding activity of c-Rel required CNBP. Lastly, -deficient mice were more susceptible to acute toxoplasmosis associated with reduced production of IL-12β, as well as a reduced T helper type 1 (Th1) cell IFN-γ response essential to controlling parasite replication. Collectively, these findings identify CNBP as important regulator of c-Rel-dependent IL-12β gene transcription and Th1 immunity.
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http://dx.doi.org/10.1084/jem.20181031DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6279399PMC
December 2018

Genetic Models Reveal cis and trans Immune-Regulatory Activities for lincRNA-Cox2.

Cell Rep 2018 11;25(6):1511-1524.e6

Department of Molecular, Cell and Developmental Biology, University of California, Santa Cruz, Santa Cruz, CA, USA. Electronic address:

An inducible gene expression program is a hallmark of the host inflammatory response. Recently, long intergenic non-coding RNAs (lincRNAs) have been shown to regulate the magnitude, duration, and resolution of these responses. Among these is lincRNA-Cox2, a dynamically regulated gene that broadly controls immune gene expression. To evaluate the in vivo functions of this lincRNA, we characterized multiple models of lincRNA-Cox2-deficient mice. LincRNA-Cox2-deficient macrophages and murine tissues had altered expression of inflammatory genes. Transcriptomic studies from various tissues revealed that deletion of the lincRNA-Cox2 locus also strongly impaired the basal and inducible expression of the neighboring gene prostaglandin-endoperoxide synthase (Ptgs2), encoding cyclooxygenase-2, a key enzyme in the prostaglandin biosynthesis pathway. By utilizing different genetic manipulations in vitro and in vivo, we found that lincRNA-Cox2 functions through an enhancer RNA mechanism to regulate Ptgs2. More importantly, lincRNA-Cox2 also functions in trans, independently of Ptgs2, to regulate critical innate immune genes in vivo.
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http://dx.doi.org/10.1016/j.celrep.2018.10.027DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6291222PMC
November 2018

Pathogen blockade of TAK1 triggers caspase-8-dependent cleavage of gasdermin D and cell death.

Science 2018 11 25;362(6418):1064-1069. Epub 2018 Oct 25.

Program in Innate Immunity, Department of Medicine, Division of Infectious Diseases and Immunology, University of Massachusetts Medical School, Worcester, MA 01605, USA.

Limited proteolysis of gasdermin D (GSDMD) generates an N-terminal pore-forming fragment that controls pyroptosis in macrophages. GSDMD is processed via inflammasome-activated caspase-1 or -11. It is currently unknown whether macrophage GSDMD can be processed by other mechanisms. Here, we describe an additional pathway controlling GSDMD processing. The inhibition of TAK1 or IκB kinase (IKK) by the effector protein YopJ elicits RIPK1- and caspase-8-dependent cleavage of GSDMD, which subsequently results in cell death. GSDMD processing also contributes to the NLRP3 inflammasome-dependent release of interleukin-1β (IL-1β). Thus, caspase-8 acts as a regulator of GSDMD-driven cell death. Furthermore, this study establishes the importance of TAK1 and IKK activity in the control of GSDMD cleavage and cytotoxicity.
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http://dx.doi.org/10.1126/science.aau2818DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6522129PMC
November 2018

Gasdermin D Restrains Type I Interferon Response to Cytosolic DNA by Disrupting Ionic Homeostasis.

Immunity 2018 09 28;49(3):413-426.e5. Epub 2018 Aug 28.

Department of Immunology, UConn Health School of Medicine, 263 Farmington Ave., Farmington, CT 06030, USA. Electronic address:

Inflammasome-activated caspase-1 cleaves gasdermin D to unmask its pore-forming activity, the predominant consequence of which is pyroptosis. Here, we report an additional biological role for gasdermin D in limiting cytosolic DNA surveillance. Cytosolic DNA is sensed by Aim2 and cyclic GMP-AMP synthase (cGAS) leading to inflammasome and type I interferon responses, respectively. We found that gasdermin D activated by the Aim2 inflammasome suppressed cGAS-driven type I interferon response to cytosolic DNA and Francisella novicida in macrophages. Similarly, interferon-β (IFN-β) response to F. novicida infection was elevated in gasdermin D-deficient mice. Gasdermin D-mediated negative regulation of IFN-β occurred in a pyroptosis-, interleukin-1 (IL-1)-, and IL-18-independent manner. Mechanistically, gasdermin D depleted intracellular potassium (K) via membrane pores, and this K efflux was necessary and sufficient to inhibit cGAS-dependent IFN-β response. Thus, our findings have uncovered an additional interferon regulatory module involving gasdermin D and K efflux.
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http://dx.doi.org/10.1016/j.immuni.2018.07.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6347470PMC
September 2018

Nrf2 negatively regulates STING indicating a link between antiviral sensing and metabolic reprogramming.

Nat Commun 2018 08 29;9(1):3506. Epub 2018 Aug 29.

Department of Biomedicine, Aarhus Research Center for Innate Immunology, Aarhus University, Bartholin Alle 6, 8000, Aarhus, Denmark.

The transcription factor Nrf2 is a critical regulator of inflammatory responses. If and how Nrf2 also affects cytosolic nucleic acid sensing is currently unknown. Here we identify Nrf2 as an important negative regulator of STING and suggest a link between metabolic reprogramming and antiviral cytosolic DNA sensing in human cells. Here, Nrf2 activation decreases STING expression and responsiveness to STING agonists while increasing susceptibility to infection with DNA viruses. Mechanistically, Nrf2 regulates STING expression by decreasing STING mRNA stability. Repression of STING by Nrf2 occurs in metabolically reprogrammed cells following TLR4/7 engagement, and is inducible by a cell-permeable derivative of the TCA-cycle-derived metabolite itaconate (4-octyl-itaconate, 4-OI). Additionally, engagement of this pathway by 4-OI or the Nrf2 inducer sulforaphane is sufficient to repress STING expression and type I IFN production in cells from patients with STING-dependent interferonopathies. We propose Nrf2 inducers as a future treatment option in STING-dependent inflammatory diseases.
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http://dx.doi.org/10.1038/s41467-018-05861-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6115435PMC
August 2018