Juan Zhang

Bio: Juan Zhang is an academic researcher from Genentech. The author has contributed to research in topics: Inflammation & Caspase-11. The author has an hindex of 21, co-authored 31 publications receiving 6237 citations.

Caspase-11 cleaves gasdermin D for non-canonical inflammasome signalling

Abstract: Intracellular lipopolysaccharide from Gram-negative bacteria including Escherichia coli, Salmonella typhimurium, Shigella flexneri, and Burkholderia thailandensis activates mouse caspase-11, causing pyroptotic cell death, interleukin-1β processing, and lethal septic shock. How caspase-11 executes these downstream signalling events is largely unknown. Here we show that gasdermin D is essential for caspase-11-dependent pyroptosis and interleukin-1β maturation. A forward genetic screen with ethyl-N-nitrosourea-mutagenized mice links Gsdmd to the intracellular lipopolysaccharide response. Macrophages from Gsdmd(-/-) mice generated by gene targeting also exhibit defective pyroptosis and interleukin-1β secretion induced by cytoplasmic lipopolysaccharide or Gram-negative bacteria. In addition, Gsdmd(-/-) mice are protected from a lethal dose of lipopolysaccharide. Mechanistically, caspase-11 cleaves gasdermin D, and the resulting amino-terminal fragment promotes both pyroptosis and NLRP3-dependent activation of caspase-1 in a cell-intrinsic manner. Our data identify gasdermin D as a critical target of caspase-11 and a key mediator of the host response against Gram-negative bacteria.

Non-canonical inflammasome activation targets caspase-11

Abstract: Caspase-1 activation by inflammasome scaffolds comprised of intracellular nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) and the adaptor ASC is believed to be essential for production of the pro-inflammatory cytokines interleukin (IL)-1β and IL-18 during the innate immune response. Here we show, with C57BL/6 Casp11 gene-targeted mice, that caspase-11 (also known as caspase-4) is critical for caspase-1 activation and IL-1β production in macrophages infected with Escherichia coli, Citrobacter rodentium or Vibrio cholerae. Strain 129 mice, like Casp11(-/-) mice, exhibited defects in IL-1β production and harboured a mutation in the Casp11 locus that attenuated caspase-11 expression. This finding is important because published targeting of the Casp1 gene was done using strain 129 embryonic stem cells. Casp1 and Casp11 are too close in the genome to be segregated by recombination; consequently, the published Casp1(-/-) mice lack both caspase-11 and caspase-1. Interestingly, Casp11(-/-) macrophages secreted IL-1β normally in response to ATP and monosodium urate, indicating that caspase-11 is engaged by a non-canonical inflammasome. Casp1(-/-)Casp11(129mt/129mt) macrophages expressing caspase-11 from a C57BL/6 bacterial artificial chromosome transgene failed to secrete IL-1β regardless of stimulus, confirming an essential role for caspase-1 in IL-1β production. Caspase-11 rather than caspase-1, however, was required for non-canonical inflammasome-triggered macrophage cell death, indicating that caspase-11 orchestrates both caspase-1-dependent and -independent outputs. Caspase-1 activation by non-canonical stimuli required NLRP3 and ASC, but caspase-11 processing and cell death did not, implying that there is a distinct activator of caspase-11. Lastly, loss of caspase-11 rather than caspase-1 protected mice from a lethal dose of lipopolysaccharide. These data highlight a unique pro-inflammatory role for caspase-11 in the innate immune response to clinically significant bacterial infections.

Noncanonical Inflammasome Activation by Intracellular LPS Independent of TLR4

Abstract: Gram-negative bacteria including Escherichia coli, Citrobacter rodentium, Salmonella typhimurium, and Shigella flexneri are sensed in an ill-defined manner by an intracellular inflammasome complex that activates caspase-11. We show that macrophages loaded with synthetic lipid A, E. coli lipopolysaccharide (LPS), or S. typhimurium LPS activate caspase-11 independently of the LPS receptor Toll-like receptor 4 (TLR4). Consistent with lipid A triggering the noncanonical inflammasome, LPS containing a divergent lipid A structure antagonized caspase-11 activation in response to E. coli LPS or Gram-negative bacteria. Moreover, LPS-mutant E. coli failed to activate caspase-11. Tlr4(-/-) mice primed with TLR3 agonist polyinosinic:polycytidylic acid [poly(I:C)] to induce pro-caspase-11 expression were as susceptible as wild-type mice were to sepsis induced by E. coli LPS. These data unveil a TLR4-independent mechanism for innate immune recognition of LPS.

Interleukin-22 alleviates metabolic disorders and restores mucosal immunity in diabetes

Abstract: In pre-clinical mouse models, the interleukin-22 pathway is identified as a novel target for therapeutic intervention in metabolic diseases.

RIPK3 deficiency or catalytically inactive RIPK1 provides greater benefit than MLKL deficiency in mouse models of inflammation and tissue injury.

Abstract: Necroptosis is a caspase-independent form of cell death that is triggered by activation of the receptor interacting serine/threonine kinase 3 (RIPK3) and phosphorylation of its pseudokinase substrate mixed lineage kinase-like (MLKL), which then translocates to membranes and promotes cell lysis. Activation of RIPK3 is regulated by the kinase RIPK1. Here we analyze the contribution of RIPK1, RIPK3, or MLKL to several mouse disease models. Loss of RIPK3 had no effect on lipopolysaccharide-induced sepsis, dextran sodium sulfate-induced colitis, cerulein-induced pancreatitis, hypoxia-induced cerebral edema, or the major cerebral artery occlusion stroke model. However, kidney ischemia–reperfusion injury, myocardial infarction, and systemic inflammation associated with A20 deficiency or high-dose tumor necrosis factor (TNF) were ameliorated by RIPK3 deficiency. Catalytically inactive RIPK1 was also beneficial in the kidney ischemia–reperfusion injury model, the high-dose TNF model, and in A20−/− mice. Interestingly, MLKL deficiency offered less protection in the kidney ischemia–reperfusion injury model and no benefit in A20−/− mice, consistent with necroptosis-independent functions for RIPK1 and RIPK3. Combined loss of RIPK3 (or MLKL) and caspase-8 largely prevented the cytokine storm, hypothermia, and morbidity induced by TNF, suggesting that the triggering event in this model is a combination of apoptosis and necroptosis. Tissue-specific RIPK3 deletion identified intestinal epithelial cells as the major target organ. Together these data emphasize that MLKL deficiency rather than RIPK1 inactivation or RIPK3 deficiency must be examined to implicate a role for necroptosis in disease.

The M1 and M2 paradigm of macrophage activation: time for reassessment.

Abstract: Macrophages are endowed with a variety of receptors for lineage-determining growth factors, T helper (Th) cell cytokines, and B cell, host, and microbial products. In tissues, macrophages mature and are activated in a dynamic response to combinations of these stimuli to acquire specialized functional phenotypes. As for the lymphocyte system, a dichotomy has been proposed for macrophage activation: classic vs. alternative, also M1 and M2, respectively. In view of recent research about macrophage functions and the increasing number of immune-relevant ligands, a revision of the model is needed. Here, we assess how cytokines and pathogen signals influence their functional phenotypes and the evidence for M1 and M2 functions and revisit a paradigm initially based on the role of a restricted set of selected ligands in the immune response.

Cleavage of GSDMD by inflammatory caspases determines pyroptotic cell death

Abstract: Inflammatory caspases (caspase-1, -4, -5 and -11) are critical for innate defences. Caspase-1 is activated by ligands of various canonical inflammasomes, and caspase-4, -5 and -11 directly recognize bacterial lipopolysaccharide, both of which trigger pyroptosis. Despite the crucial role in immunity and endotoxic shock, the mechanism for pyroptosis induction by inflammatory caspases is unknown. Here we identify gasdermin D (Gsdmd) by genome-wide clustered regularly interspaced palindromic repeat (CRISPR)-Cas9 nuclease screens of caspase-11- and caspase-1-mediated pyroptosis in mouse bone marrow macrophages. GSDMD-deficient cells resisted the induction of pyroptosis by cytosolic lipopolysaccharide and known canonical inflammasome ligands. Interleukin-1β release was also diminished in Gsdmd(-/-) cells, despite intact processing by caspase-1. Caspase-1 and caspase-4/5/11 specifically cleaved the linker between the amino-terminal gasdermin-N and carboxy-terminal gasdermin-C domains in GSDMD, which was required and sufficient for pyroptosis. The cleavage released the intramolecular inhibition on the gasdermin-N domain that showed intrinsic pyroptosis-inducing activity. Other gasdermin family members were not cleaved by inflammatory caspases but shared the autoinhibition; gain-of-function mutations in Gsdma3 that cause alopecia and skin defects disrupted the autoinhibition, allowing its gasdermin-N domain to trigger pyroptosis. These findings offer insight into inflammasome-mediated immunity/diseases and also change our understanding of pyroptosis and programmed necrosis.

Molecular mechanisms of cell death: recommendations of the Nomenclature Committee on Cell Death 2018.

Abstract: Over the past decade, the Nomenclature Committee on Cell Death (NCCD) has formulated guidelines for the definition and interpretation of cell death from morphological, biochemical, and functional perspectives. Since the field continues to expand and novel mechanisms that orchestrate multiple cell death pathways are unveiled, we propose an updated classification of cell death subroutines focusing on mechanistic and essential (as opposed to correlative and dispensable) aspects of the process. As we provide molecularly oriented definitions of terms including intrinsic apoptosis, extrinsic apoptosis, mitochondrial permeability transition (MPT)-driven necrosis, necroptosis, ferroptosis, pyroptosis, parthanatos, entotic cell death, NETotic cell death, lysosome-dependent cell death, autophagy-dependent cell death, immunogenic cell death, cellular senescence, and mitotic catastrophe, we discuss the utility of neologisms that refer to highly specialized instances of these processes. The mission of the NCCD is to provide a widely accepted nomenclature on cell death in support of the continued development of the field.

Activation and regulation of the inflammasomes

Abstract: Inflammasomes are key signalling platforms that detect pathogenic microorganisms and sterile stressors, and that activate the highly pro-inflammatory cytokines interleukin-1β (IL-1β) and IL-18. In this Review, we discuss the complex regulatory mechanisms that facilitate a balanced but effective inflammasome-mediated immune response, and we highlight the similarities to another molecular signalling platform - the apoptosome - that monitors cellular health. Extracellular regulatory mechanisms are discussed, as well as the intracellular control of inflammasome assembly, for example, via ion fluxes, free radicals and autophagy.