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1.
J Exp Med ; 221(8)2024 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-38780621

RESUMO

Nucleic acid-sensing Toll-like receptors (TLR) 3, 7/8, and 9 are key innate immune sensors whose activities must be tightly regulated to prevent systemic autoimmune or autoinflammatory disease or virus-associated immunopathology. Here, we report a systematic scanning-alanine mutagenesis screen of all cytosolic and luminal residues of the TLR chaperone protein UNC93B1, which identified both negative and positive regulatory regions affecting TLR3, TLR7, and TLR9 responses. We subsequently identified two families harboring heterozygous coding mutations in UNC93B1, UNC93B1+/T93I and UNC93B1+/R336C, both in key negative regulatory regions identified in our screen. These patients presented with cutaneous tumid lupus and juvenile idiopathic arthritis plus neuroinflammatory disease, respectively. Disruption of UNC93B1-mediated regulation by these mutations led to enhanced TLR7/8 responses, and both variants resulted in systemic autoimmune or inflammatory disease when introduced into mice via genome editing. Altogether, our results implicate the UNC93B1-TLR7/8 axis in human monogenic autoimmune diseases and provide a functional resource to assess the impact of yet-to-be-reported UNC93B1 mutations.


Assuntos
Autoimunidade , Animais , Humanos , Camundongos , Autoimunidade/genética , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Análise Mutacional de DNA , Receptores Toll-Like/metabolismo , Receptores Toll-Like/genética , Mutação , Feminino , Masculino , Camundongos Endogâmicos C57BL , Células HEK293 , Receptor 7 Toll-Like/genética , Receptor 7 Toll-Like/metabolismo , Doenças Autoimunes/genética , Doenças Autoimunes/imunologia
2.
Elife ; 122023 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-37417868

RESUMO

Inflammasomes are cytosolic innate immune complexes that assemble upon detection of diverse pathogen-associated cues and play a critical role in host defense and inflammatory pathogenesis. Here, we find that the human inflammasome-forming sensor CARD8 senses HIV-1 infection via site-specific cleavage of the CARD8 N-terminus by the HIV protease (HIV-1PR). HIV-1PR cleavage of CARD8 induces pyroptotic cell death and the release of pro-inflammatory cytokines from infected cells, processes regulated by Toll-like receptor stimulation prior to viral infection. In acutely infected cells, CARD8 senses the activity of both de novo translated HIV-1PR and packaged HIV-1PR that is released from the incoming virion. Moreover, our evolutionary analyses reveal that the HIV-1PR cleavage site in human CARD8 arose after the divergence of chimpanzees and humans. Although chimpanzee CARD8 does not recognize proteases from HIV or simian immunodeficiency viruses from chimpanzees (SIVcpz), SIVcpz does cleave human CARD8, suggesting that SIVcpz was poised to activate the human CARD8 inflammasome prior to its cross-species transmission into humans. Our findings suggest a unique role for CARD8 inflammasome activation in response to lentiviral infection of humans.


Assuntos
Infecções por HIV , HIV-1 , Vírus da Imunodeficiência Símia , Animais , Humanos , Inflamassomos/metabolismo , Pan troglodytes/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Proteínas de Neoplasias/metabolismo , Proteínas Adaptadoras de Sinalização CARD/metabolismo
3.
PLoS Biol ; 21(6): e3002144, 2023 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-37289745

RESUMO

Hosts have evolved diverse strategies to respond to microbial infections, including the detection of pathogen-encoded proteases by inflammasome-forming sensors such as NLRP1 and CARD8. Here, we find that the 3CL protease (3CLpro) encoded by diverse coronaviruses, including Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), cleaves a rapidly evolving region of human CARD8 and activates a robust inflammasome response. CARD8 is required for cell death and the release of pro-inflammatory cytokines during SARS-CoV-2 infection. We further find that natural variation alters CARD8 sensing of 3CLpro, including 3CLpro-mediated antagonism rather than activation of megabat CARD8. Likewise, we find that a single nucleotide polymorphism (SNP) in humans reduces CARD8's ability to sense coronavirus 3CLpros and, instead, enables sensing of 3C proteases (3Cpro) from select picornaviruses. Our findings demonstrate that CARD8 is a broad sensor of viral protease activities and suggests that CARD8 diversity contributes to inter- and intraspecies variation in inflammasome-mediated viral sensing and immunopathology.


Assuntos
COVID-19 , Picornaviridae , Humanos , Inflamassomos/metabolismo , Picornaviridae/genética , Picornaviridae/metabolismo , SARS-CoV-2/metabolismo , Inibidores de Proteases , Proteínas Reguladoras de Apoptose/metabolismo , Proteínas de Neoplasias/metabolismo , Proteínas Adaptadoras de Sinalização CARD/metabolismo
4.
Elife ; 122023 01 16.
Artigo em Inglês | MEDLINE | ID: mdl-36645406

RESUMO

Bacteria of the genus Shigella cause shigellosis, a severe gastrointestinal disease driven by bacterial colonization of colonic intestinal epithelial cells. Vertebrates have evolved programmed cell death pathways that sense invasive enteric pathogens and eliminate their intracellular niche. Previously we reported that genetic removal of one such pathway, the NAIP-NLRC4 inflammasome, is sufficient to convert mice from resistant to susceptible to oral Shigella flexneri challenge (Mitchell et al., 2020). Here, we investigate the protective role of additional cell death pathways during oral mouse Shigella infection. We find that the Caspase-11 inflammasome, which senses Shigella LPS, restricts Shigella colonization of the intestinal epithelium in the absence of NAIP-NLRC4. However, this protection is limited when Shigella expresses OspC3, an effector that antagonizes Caspase-11 activity. TNFα, a cytokine that activates Caspase-8-dependent apoptosis, also provides potent protection from Shigella colonization of the intestinal epithelium when mice lack both NAIP-NLRC4 and Caspase-11. The combined genetic removal of Caspases-1, -11, and -8 renders mice hyper-susceptible to oral Shigella infection. Our findings uncover a layered hierarchy of cell death pathways that limit the ability of an invasive gastrointestinal pathogen to cause disease.


Assuntos
Disenteria Bacilar , Shigella , Camundongos , Animais , Disenteria Bacilar/microbiologia , Inflamassomos/metabolismo , Morte Celular , Shigella flexneri/metabolismo , Caspases/genética , Caspases/metabolismo
5.
bioRxiv ; 2022 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-36172130

RESUMO

Hosts have evolved diverse strategies to respond to microbial infections, including the detection of pathogen-encoded proteases by inflammasome-forming sensors such as NLRP1 and CARD8. Here, we find that the 3CL protease (3CL pro ) encoded by diverse coronaviruses, including SARS-CoV-2, cleaves a rapidly evolving region of human CARD8 and activates a robust inflammasome response. CARD8 is required for cell death and the release of pro-inflammatory cytokines during SARS-CoV-2 infection. We further find that natural variation alters CARD8 sensing of 3CL pro , including 3CL pro -mediated antagonism rather than activation of megabat CARD8. Likewise, we find that a single nucleotide polymorphism (SNP) in humans reduces CARD8’s ability to sense coronavirus 3CL pros , and instead enables sensing of 3C proteases (3C pro ) from select picornaviruses. Our findings demonstrate that CARD8 is a broad sensor of viral protease activities and suggests that CARD8 diversity contributes to inter- and intra-species variation in inflammasome-mediated viral sensing and immunopathology.

6.
Proc Natl Acad Sci U S A ; 119(7)2022 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-35135876

RESUMO

Gasdermins are a family of pore-forming proteins controlling an inflammatory cell death reaction in the mammalian immune system. The pore-forming ability of the gasdermin proteins is released by proteolytic cleavage with the removal of their inhibitory C-terminal domain. Recently, gasdermin-like proteins have been discovered in fungi and characterized as cell death-inducing toxins in the context of conspecific non-self-discrimination (allorecognition). Although functional analogies have been established between mammalian and fungal gasdermins, the molecular pathways regulating gasdermin activity in fungi remain largely unknown. Here, we characterize a gasdermin-based cell death reaction controlled by the het-Q allorecognition genes in the filamentous fungus Podospora anserina We show that the cytotoxic activity of the HET-Q1 gasdermin is controlled by proteolysis. HET-Q1 loses a ∼5-kDa C-terminal fragment during the cell death reaction in the presence of a subtilisin-like serine protease termed HET-Q2. Mutational analyses and successful reconstitution of the cell death reaction in heterologous hosts (Saccharomyces cerevisiae and human 293T cells) suggest that HET-Q2 directly cleaves HET-Q1 to induce cell death. By analyzing the genomic landscape of het-Q1 homologs in fungi, we uncovered that the vast majority of the gasdermin genes are clustered with protease-encoding genes. These HET-Q2-like proteins carry either subtilisin-like or caspase-related proteases, which, in some cases, correspond to the N-terminal effector domain of nucleotide-binding and oligomerization-like receptor proteins. This study thus reveals the proteolytic regulation of gasdermins in fungi and establishes evolutionary parallels between fungal and mammalian gasdermin-dependent cell death pathways.


Assuntos
Proteínas Fúngicas/metabolismo , Regulação Fúngica da Expressão Gênica/fisiologia , Podospora/metabolismo , Apoptose/fisiologia , Morte Celular , Sobrevivência Celular , Proteínas Fúngicas/genética , Células HEK293 , Humanos , Podospora/genética , Proteólise , Saccharomyces cerevisiae , Subtilisina
7.
PLoS One ; 15(11): e0241922, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33186363

RESUMO

Inertial measurement units (IMUs) have been increasingly popular in rehabilitation research. However, despite their accessibility and potential advantages, their uptake and acceptance by health professionals remain a big challenge. The development of an IMU-based clinical tool must bring together engineers, researchers and clinicians. This study is part of a developmental process with the investigation of clinicians' perspectives about IMUs. Clinicians from four rehabilitation centers were invited to a 30-minute presentation on IMUs. Then, two one-hour focus groups were conducted with volunteer clinicians in each rehabilitation center on: 1) IMUs and their clinical usefulness, and 2) IMUs data analysis and visualization interface. Fifteen clinicians took part in the first focus groups. They expressed their thoughts on: 1) categories of variables that would be useful to measure with IMUs in clinical practice, and 2) desired characteristics of the IMUs. Twenty-three clinicians participated to the second focus groups, discussing: 1) functionalities, 2) display options, 3) clinical data reported and associated information, and 4) data collection duration. Potential influence of IMUs on clinical practice and added value were discussed in both focus groups. Clinicians expressed positive opinions about the use of IMUs, but their expectations were high before considering using IMUs in their practice.


Assuntos
Médicos/psicologia , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Centros de Reabilitação , Adulto Jovem
8.
Elife ; 92020 10 19.
Artigo em Inglês | MEDLINE | ID: mdl-33074100

RESUMO

Bacteria of the genus Shigella cause shigellosis, a severe gastrointestinal disease that is a major cause of diarrhea-associated mortality in humans. Mice are highly resistant to Shigella and the lack of a tractable physiological model of shigellosis has impeded our understanding of this important human disease. Here, we propose that the differential susceptibility of mice and humans to Shigella is due to mouse-specific activation of the NAIP-NLRC4 inflammasome. We find that NAIP-NLRC4-deficient mice are highly susceptible to oral Shigella infection and recapitulate the clinical features of human shigellosis. Although inflammasomes are generally thought to promote Shigella pathogenesis, we instead demonstrate that intestinal epithelial cell (IEC)-specific NAIP-NLRC4 activity is sufficient to protect mice from shigellosis. In addition to describing a new mouse model of shigellosis, our results suggest that the lack of an inflammasome response in IECs may help explain the susceptibility of humans to shigellosis.


Assuntos
Proteínas Reguladoras de Apoptose/deficiência , Proteínas de Ligação ao Cálcio/deficiência , Suscetibilidade a Doenças/imunologia , Disenteria Bacilar/imunologia , Proteína Inibidora de Apoptose Neuronal/deficiência , Animais , Humanos , Inflamassomos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Shigella/imunologia
9.
J Therm Biol ; 93: 102701, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-33077122

RESUMO

In some turtle species, temperature selection may be influenced by environmental conditions, including acclimation temperature and substrate quality. These factors may be particularly important for softshell turtles that are highly aquatic and often thermoregulate by burying in the substrate in shallow water microhabitats. We tested for effects of acclimation temperature (22 °C or 27 °C) and substrate type (sand or gravel) on the selected temperature and movement patterns of 20 juvenile spiny softhshell turtles (Apalone spinifera; Reptilia: Trionychidae) in an aquatic thermal gradient of 14-34 °C. Among 7-11 month old juvenile softshell turtles, acclimation temperature and substrate type did not influence temperature selection, nor alter activity and movement patterns. During thermal gradient tests, both 22- and 27 °C-acclimated turtles selected the warmest temperature (34 °C) available most frequently, regardless of substrate type (sand or gravel). Similarly, acclimation temperature and substrate type did not influence movement patterns of turtles, nor the number of chambers used in the gradient tests. These results suggest that juvenile Apalone spinifera are capable of detecting small temperature increments and prefer warm temperatures that may positively influence growth and metabolism, and that thermal factors more significantly influence aquatic thermoregulation in this species than does substrate type.


Assuntos
Aclimatação , Distribuição Animal , Temperatura , Tartarugas/fisiologia , Animais , Movimento , Areia , Tartarugas/crescimento & desenvolvimento
11.
Am J Trop Med Hyg ; 98(1): 142-145, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-29165220

RESUMO

Leishmania infantum causes visceral leishmaniasis (VL) in Brazil. We previously observed that VL is more common in males than females living in endemic neighborhoods, despite similar exposure. Using a larger sample, we document that VL is more common in males than females, but only after puberty. BALB/c and C57BL/6 mouse models confirmed that there is a biological basis for male susceptibility to symptomatic VL, showing higher parasite burdens in males than females. Female C57BL/6 mice generated more antigen-induced cytokines associated with curative responses (interferon-γ, interleukin [IL]-1ß). Males expressed higher levels of IL-10 and tumor necrosis factor, which are linked to exacerbated disease. Different parasite lines entered or survived at a higher rate in macrophages of male- than female-origin. These results suggest that males are inherently more susceptible to L. infantum than females and that mice are a valid model to study this sex-dependent difference.


Assuntos
Leishmania infantum , Leishmaniose Visceral/epidemiologia , Adolescente , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Animais , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Interferon gama/sangue , Interleucina-10/sangue , Interleucina-1beta/sangue , Leishmaniose Visceral/tratamento farmacológico , Leishmaniose Visceral/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Fatores Sexuais , África do Sul/epidemiologia , Fator de Necrose Tumoral alfa/sangue , Adulto Jovem
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