38 results found
Fisch D, Bando H, Clough B, et al., 2019, Human GBP1 is a microbe-specific gatekeeper of macrophage apoptosis and pyroptosis., EMBO J
The guanylate binding protein (GBP) family of interferon-inducible GTPases promotes antimicrobial immunity and cell death. During bacterial infection, multiple mouse Gbps, human GBP2, and GBP5 support the activation of caspase-1-containing inflammasome complexes or caspase-4 which trigger pyroptosis. Whether GBPs regulate other forms of cell death is not known. The apicomplexan parasite Toxoplasma gondii causes macrophage death through unidentified mechanisms. Here we report that Toxoplasma-induced death of human macrophages requires GBP1 and its ability to target Toxoplasma parasitophorous vacuoles through its GTPase activity and prenylation. Mechanistically, GBP1 promoted Toxoplasma detection by AIM2, which induced GSDMD-independent, ASC-, and caspase-8-dependent apoptosis. Identical molecular determinants targeted GBP1 to Salmonella-containing vacuoles. GBP1 facilitated caspase-4 recruitment to Salmonella leading to its enhanced activation and pyroptosis. Notably, GBP1 could be bypassed by the delivery of Toxoplasma DNA or bacterial LPS into the cytosol, pointing to its role in liberating microbial molecules. GBP1 thus acts as a gatekeeper of cell death pathways, which respond specifically to infecting microbes. Our findings expand the immune roles of human GBPs in regulating not only pyroptosis, but also apoptosis.
Goddard P, Sanchez Garrido J, Slater S, et al., 2019, Enteropathogenic E. coli stimulates effector-driven rapid caspase-4 activation in human macrophages, Cell Reports, Vol: 27, Pages: 1008-1017.e6, ISSN: 2211-1247
Microbial infections can stimulate the assembly of inflammasomes, which activate caspase-1. The gastrointestinal pathogen enteropathogenic Escherichia coli (EPEC) causes localized actin polymerization in host cells. Actin polymerization requires the binding of the bacterial adhesin intimin to Tir, which is delivered to host cells via a type 3 secretion system (T3SS). We show that EPEC induces T3SS-dependent rapid non-canonical NLRP3 inflammasome activation in human macrophages. Notably, caspase-4 activation by EPEC triggers pyroptosis and cytokine processing through the NLRP3-caspase-1 inflammasome. Mechanistically, caspase-4 activation requires the detection of LPS and EPEC-induced actin polymerization, either via Tir tyrosine phosphorylation and the phosphotyrosine-binding adaptor NCK or Tir and the NCK-mimicking effector TccP. An engineered E. coli K12 could reconstitute Tir-intimin signaling, which is necessary and sufficient for inflammasome activation, ruling out the involvement of other virulence factors. Our studies reveal a crosstalk between caspase-4 and caspase-1 that is cooperatively stimulated by LPS and effector-driven actin polymerization.
Sanchez-Garrdio J, Sancho-Shimizu V, Shenoy A, 2018, Regulated proteolysis of p62/SQSTM1 enables differential control of autophagy and nutrient sensing, Science Signaling, Vol: 11, ISSN: 1937-9145
The multidomain scaffold protein p62 (also called sequestosome-1) is involved in autophagy, antimicrobial immunity, and oncogenesis. Mutations in SQSTM1, which encodes p62, are linked to hereditary inflammatory conditions such as Paget’s disease of the bone, frontotemporal dementia (FTD), amyotrophic lateral sclerosis, and distal myopathy with rimmed vacuoles. Here, we report that p62 was proteolytically trimmed by the protease caspase-8 into a stable protein, which we called p62TRM. We found that p62TRM, but not full-length p62, was involved in nutrient sensing and homeostasis through the mechanistic target of rapamycin complex 1 (mTORC1). The kinase RIPK1 and caspase-8 controlled p62TRM production and thus promoted mTORC1 signaling. An FTD-linked p62 D329G polymorphism and a rare D329H variant could not be proteolyzed by caspase-8, and these noncleavable variants failed to activate mTORC1, thereby revealing the detrimental effect of these mutations. These findings on the role of p62TRM provide new insights into SQSTM1-linked diseases and mTORC1 signaling.
Ahmad L, Mashbat B, Leung C, et al., 2018, Human TANK-binding kinase 1 is required for early autophagy induction upon herpes simplex virus 1 infection, Journal of Allergy and Clinical Immunology, ISSN: 0091-6749
Shenoy AR, Furniss RCD, Goddard PJ, et al., 2018, Modulation of host cell processes by T3SS effectors, Publisher: Springer Verlag
Two of the enteric Escherichia coli pathotypes-enteropathogenic E. coli (EPEC) and enterohaemorrhagic E. coli (EHEC)-have a conserved type 3 secretion system which is essential for virulence. The T3SS is used to translocate between 25 and 50 bacterial proteins directly into the host cytosol where they manipulate a variety of host cell processes to establish a successful infection. In this chapter, we discuss effectors from EPEC/EHEC in the context of the host proteins and processes that they target-the actin cytoskeleton, small guanosine triphosphatases and innate immune signalling pathways that regulate inflammation and cell death. Many of these translocated proteins have been extensively characterised, which has helped obtain insights into the mechanisms of pathogenesis of these bacteria and also understand the host pathways they target in more detail. With increasing knowledge of the positive and negative regulation of host signalling pathways by different effectors, a future challenge is to investigate how the specific effector repertoire of each strain cooperates over the course of an infection.
Pallett MA, Crepin VF, Serafini N, et al., 2017, Bacterial Virulence Factor Inhibits Caspase-4/11 Activation in IntestinalEpithelial Cells, Mucosal Immunology, Vol: 10, Pages: 602-612, ISSN: 1935-3456
The human pathogen enteropathogenic Escherichia coli (EPEC), as well as the mouse pathogen Citrobacter rodentium, colonize the gut mucosa via attaching and effacing lesion formation and cause diarrheal diseases. EPEC and C. rodentium type III secretion system (T3SS) effectors repress innate immune responses and infiltration of immune cells. Inflammatory caspases such as caspase'1 and caspase'4/11 are crucial mediators of host defense and inflammation in the gut via their ability to process cytokines such as IL'1β and IL'18. Here we report that the effector NleF binds the catalytic domain of caspase'4 and inhibits its proteolytic activity. Following infection of intestinal epithelial cells (IECs) EPEC inhibited caspase'4 and IL'18 processing in an NleF'dependent manner. Depletion of caspase'4 in IECs prevented the secretion of mature IL'18 in response to infection with EPEC@nleF. NleF'dependent inhibition of caspase'11 in colons of mice prevented IL'18 secretion and neutrophil influx at early stages of C. rodentium infection. Neither wild'type C. rodentium nor C. rodentium@nleF triggered neutrophil infiltration or IL'18 secretion in Cas11 or Casp1/11 deficient mice. Thus, IECs play a key role in modulating early innate immune responses in the gut via a caspase'4/11 ' IL'18 axis, which is targeted by virulence factors encoded by enteric pathogens
Bist P, Cheong WS, Ng A, et al., 2017, E3 Ubiquitin ligase ZNRF4 negatively regulates NOD2 signalling and induces tolerance to MDP, Nature Communications, Vol: 8, ISSN: 2041-1723
Optimal regulation of the innate immune receptor nucleotide-binding oligomerization domain-containing protein 2 (NOD2) is essential for controlling bacterial infections and inflammatory disorders. Chronic NOD2 stimulation induces non-responsiveness to restimulation, termed NOD2-induced tolerance. Although the levels of the NOD2 adaptor, RIP2, are reported to regulate both acute and chronic NOD2 signalling, how RIP2 levels are modulated is unclear. Here we show that ZNRF4 induces K48-linked ubiquitination of RIP2 and promotes RIP2 degradation. A fraction of RIP2 localizes to the endoplasmic reticulum (ER), where it interacts with ZNRF4 under either unstimulated and muramyl dipeptide-stimulated conditions. Znrf4 knockdown monocytes have sustained nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) activation, and Znrf4 knockdown mice have reduced NOD2-induced tolerance and more effective control of Listeria monocytogenes infection. Our results thus demonstrate E3-ubiquitin ligase ZNRF4-mediated RIP2 degradation as a negative regulatory mechanism of NOD2-induced NF-κB, cytokine and anti-bacterial responses in vitro and in vivo, and identify a ZNRF4-RIP2 axis of fine-tuning NOD2 signalling to promote protective host immunity.
Mazon-Moya MJ, Willis AR, Torraca V, et al., 2017, Septins restrict inflammation and protect zebrafish larvae from Shigella infection, PLoS Pathogens, Vol: 13, ISSN: 1553-7366
Shigella flexneri, a Gram-negative enteroinvasive pathogen, causes inflammatory destruction of the human intestinal epithelium. Infection by S. flexneri has been well-studied in vitro and is a paradigm for bacterial interactions with the host immune system. Recent work has revealed that components of the cytoskeleton have important functions in innate immunity and inflammation control. Septins, highly conserved cytoskeletal proteins, have emerged as key players in innate immunity to bacterial infection, yet septin function in vivo is poorly understood. Here, we use S. flexneri infection of zebrafish (Danio rerio) larvae to study in vivo the role of septins in inflammation and infection control. We found that depletion of Sept15 or Sept7b, zebrafish orthologs of human SEPT7, significantly increased host susceptibility to bacterial infection. Live-cell imaging of Sept15-depleted larvae revealed increasing bacterial burdens and a failure of neutrophils to control infection. Strikingly, Sept15-depleted larvae present significantly increased activity of Caspase-1 and more cell death upon S. flexneri infection. Dampening of the inflammatory response with anakinra, an antagonist of interleukin-1 receptor (IL-1R), counteracts Sept15 deficiency in vivo by protecting zebrafish from hyper-inflammation and S. flexneri infection. These findings highlight a new role for septins in host defence against bacterial infection, and suggest that septin dysfunction may be an underlying factor in cases of hyper-inflammation.
Eldridge MJG, Sanchez Garrido J, Hoben GF, et al., 2017, The atypical ubiquitin E2 conjugase UBE2L3 is an indirect caspase-1 target and controls IL-1beta secretion by inflammasomes, Cell Reports, Vol: 18, Pages: 1285-1297, ISSN: 2211-1247
Caspase-1 activation by inflammasome signalling scaffoldsinitiates inflammation and antimicrobial responses. Caspase-1 proteolytically converts newly induced pro-IL-1α into its mature form and directs its secretion, triggers pyroptosis and the release of non-substrate alarmins such as IL-1α and HMGB1. While somecaspase-1 substrates involved in these events are known, the identities and roles of non-proteolytic targets remain unknown. Here we report using unbiased proteomics that the UBE2L3 ubiquitin conjugase is an indirect target of caspase-1. Caspase-1, but not caspase-4, controlled pyroptosis-and ubiquitin-independent proteasomal degradation of UBE2L3 upon canonical and non-canonical inflammasome activation by sterile danger signals and bacterial infection. Mechanistically, UBE2L3 acted post-translationally to promote K48-ubiquitylation and turnover of pro-IL-1β and dampen mature-IL-1β production. UBE2L3 depletion increased pro-IL-1β levels and mature-IL-1βsecretion by inflammasomes. These findings on UBE2L3 as a molecular rheostat have implications for IL-1-driven pathology in hereditary fever syndromes, and autoinflammatory conditions associated with UBE2L3 polymorphisms.
Thurston T, Matthews S, Jennings E, et al., 2016, Growth inhibition of cytosolic Salmonella by caspase-1 and caspase-11 precedes host cell death, Nature Communications, Vol: 7, ISSN: 2041-1723
Sensing bacterial products in the cytosol of mammalian cells by NOD-like receptors leads to the activation of caspase-1 inflammasomes, and the production of the pro-inflammatory cytokines interleukin (IL)-18 and IL-1β. In addition, mouse caspase-11 (represented in humans by its orthologs, caspase-4 and caspase-5) detects cytosolic bacterial LPS directly. Activation of caspase-1 and caspase-11 initiates pyroptotic host cell death that releases potentially harmful bacteria from the nutrient-rich host cell cytosol into the extracellular environment. Here we use single cell analysis and time-lapse microscopy to identify a subpopulation of host cells, in which growth of cytosolic Salmonella Typhimurium is inhibited independently or prior to the onset of cell death. The enzymatic activities of caspase-1 and caspase-11 are required for growth inhibition in different cell types. Our results reveal that these proteases have important functions beyond the direct induction of pyroptosis and proinflammatory cytokine secretion in the control of growth and elimination of cytosolic bacteria.
Surana S, Shenoy AR, Krishnan Y, 2015, Designing DNA nanodevices for compatibility with the immune system of higher organisms, Nature Nanotechnology, Vol: 10, Pages: 741-747, ISSN: 1748-3395
DNA is proving to be a powerful scaffold to construct molecularly precise designer DNA devices. Recent trends reveal their ever-increasing deployment within living systems as delivery devices that not only probe but also program and re-program a cell, or even whole organisms. Given that DNA is highly immunogenic, we outline the molecular, cellular and organismal response pathways that designer nucleic acid nanodevices are likely to elicit in living systems. We address safety issues applicable when such designer DNA nanodevices interact with the immune system. In light of this, we discuss possible molecular programming strategies that could be integrated with such designer nucleic acid scaffolds to either evade or stimulate the host response with a view to optimizing and widening their applications in higher organisms.
Mostowy S, Shenoy AR, 2015, The cytoskeleton in cell-autonomous immunity: structural determinants of host defence, Nature Reviews Immunology, Vol: 15, Pages: 559-573, ISSN: 1474-1741
Host cells use antimicrobial proteins, pathogen-restrictive compartmentalization and cell death in their defence against intracellular pathogens. Recent work has revealed that four components of the cytoskeleton — actin, microtubules, intermediate filaments and septins, which are well known for their roles in cell division, shape and movement — have important functions in innate immunity and cellular self-defence. Investigations using cellular and animal models have shown that these cytoskeletal proteins are crucial for sensing bacteria and for mobilizing effector mechanisms to eliminate them. In this Review, we highlight the emerging roles of the cytoskeleton as a structural determinant of cell-autonomous host defence.
Eldridge MJG, Shenoy AR, 2015, Antimicrobial inflammasomes: unified signalling against diverse bacterial pathogens, CURRENT OPINION IN MICROBIOLOGY, Vol: 23, Pages: 32-41, ISSN: 1369-5274
Matsuzawa T, Kim B-H, Shenoy AR, et al., 2012, IFN-γ elicits macrophage autophagy via the p38 MAPK signaling pathway., J Immunol, Vol: 189, Pages: 813-818
Autophagy is a major innate immune defense pathway in both plants and animals. In mammals, this cascade can be elicited by cytokines (IFN-γ) or pattern recognition receptors (TLRs and nucleotide-binding oligomerization domain-like receptors). Many signaling components in TLR- and nucleotide-binding oligomerization domain-like receptor-induced autophagy are now known; however, those involved in activating autophagy via IFN-γ remain to be elucidated. In this study, we engineered macrophages encoding a tandem fluorescently tagged LC3b (tfLC3) autophagosome reporter along with stably integrated short hairpin RNAs to demonstrate IFN-γ-induced autophagy required JAK 1/2, PI3K, and p38 MAPK but not STAT1. Moreover, the autophagy-related guanosine triphosphatase Irgm1 proved dispensable in both stable tfLC3-expressing RAW 264.7 and tfLC3-transduced Irgm1(-/-) primary macrophages, revealing a novel p38 MAPK-dependent, STAT1-independent autophagy pathway that bypasses Irgm1. These unexpected findings have implications for understanding how IFN-γ-induced autophagy is mobilized within macrophages for inflammation and host defense.
Shenoy AR, Wellington DA, Kumar P, et al., 2012, GBP5 Promotes NLRP3 Inflammasome Assembly and Immunity in Mammals, SCIENCE, Vol: 336, Pages: 481-485, ISSN: 0036-8075
Kim B-H, Shenoy AR, Kumar P, et al., 2011, A family of IFN-γ-inducible 65-kD GTPases protects against bacterial infection., Science, Vol: 332, Pages: 717-721
Immune interferon gamma (IFN-γ) is essential for mammalian host defense against intracellular pathogens. IFN-γ induces nearly 2000 host genes, yet few have any assigned function. Here, we examined a complete mouse 65-kilodalton (kD) guanylate-binding protein (Gbp) gene family as part of a 43-member IFN-γ-inducible guanosine triphosphatase (GTPase) superfamily in mouse and human genomes. Family-wide loss-of-function analysis found that at least four Gbps--Gbp1, Gbp6, Gbp7, and Gbp10--conferred cell-autonomous immunity to listerial or mycobacterial infection within macrophages and gene-deficient animals. These Gbps solicited host defense proteins, including the phagocyte oxidase, antimicrobial peptides, and autophagy effectors, to kill intracellular bacteria. Thus, specific 65-kD Gbps coordinate a potent oxidative and vesicular trafficking program to protect the host from infection.
Biswas KH, Shenoy AR, Dutta A, et al., 2009, The Evolution of Guanylyl Cyclases as Multidomain Proteins: Conserved Features of Kinase-Cyclase Domain Fusions, JOURNAL OF MOLECULAR EVOLUTION, Vol: 68, Pages: 587-602, ISSN: 0022-2844
Tyagi R, Shenoy AR, Visweswariah SS, 2009, Characterization of an Evolutionarily Conserved Metallophosphoesterase That Is Expressed in the Fetal Brain and Associated with the WAGR Syndrome, JOURNAL OF BIOLOGICAL CHEMISTRY, Vol: 284, Pages: 5217-5228, ISSN: 0021-9258
Dass BKM, Sharma R, Shenoy AR, et al., 2008, Cyclic AMP in mycobacteria: Characterization and functional role of the rv1647 ortholog in Mycobacterium smegmatis, JOURNAL OF BACTERIOLOGY, Vol: 190, Pages: 3824-3834, ISSN: 0021-9193
Shenoy AR, Capuder M, Draskovic P, et al., 2007, Structural and biochemical analysis of the Rv0805 cyclic nucleotide phosphodiesterase from Mycobacterium tuberculosis, JOURNAL OF MOLECULAR BIOLOGY, Vol: 365, Pages: 211-225, ISSN: 0022-2836
Shenoy AR, Kim B-H, Choi H-P, et al., 2007, Emerging themes in IFN-gamma-induced macrophage immunity by the p47 and p65 GTPase families, IMMUNOBIOLOGY, Vol: 212, Pages: 771-784, ISSN: 0171-2985
Shenoy AR, Visweswariah SS, 2006, New messages from old messengers: cAMP and mycobacteria, TRENDS IN MICROBIOLOGY, Vol: 14, Pages: 543-550, ISSN: 0966-842X
Macario AJL, Brocchieri L, Shenoy AR, et al., 2006, Evolution of a protein-folding machine: Genomic and evolutionary analyses reveal three lineages of the archaeal hsp70(dnaK) gene, JOURNAL OF MOLECULAR EVOLUTION, Vol: 63, Pages: 74-86, ISSN: 0022-2844
Shenoy AR, Visweswariah SS, 2006, Mycobacterial adenylyl cyclases: Biochemical diversity and structural plasticity, FEBS LETTERS, Vol: 580, Pages: 3344-3352, ISSN: 1873-3468
Ketkar AD, Shenoy AR, Ramagopal UA, et al., 2006, A structural basis for the role of nucleotide specifying residues in regulating the oligomerization of the Rv1625c adenylyl cyclase from M. tuberculosis., J Mol Biol, Vol: 356, Pages: 904-916, ISSN: 0022-2836
The Rv1625c Class III adenylyl cyclase from Mycobacterium tuberculosis is a homodimeric enzyme with two catalytic centers at the dimer interface, and shows sequence similarity with the mammalian adenylyl and guanylyl cyclases. Mutation of the substrate-specifying residues in the catalytic domain of Rv1625c, either independently or together, to those present in guanylyl cyclases not only failed to confer guanylyl cyclase activity to the protein, but also severely abrogated the adenylyl cyclase activity of the enzyme. Biochemical analysis revealed alterations in the behavior of the mutants on ion-exchange chromatography, indicating differences in the surface-exposed charge upon mutation of substrate-specifying residues. The mutant proteins showed alterations in oligomeric status as compared to the wild-type enzyme, and differing abilities to heterodimerize with the wild-type protein. The crystal structure of a mutant has been solved to a resolution of 2.7A. On the basis of the structure, and additional biochemical studies, we provide possible reasons for the altered properties of the mutant proteins, as well as highlight unique structural features of the Rv1625c adenylyl cyclase.
Jaleel M, Saha S, Shenoy AR, et al., 2006, The kinase homology domain of receptor guanylyl cyclase C: ATP binding and identification of an adenine nucleotide sensitive site, BIOCHEMISTRY, Vol: 45, Pages: 1888-1898, ISSN: 0006-2960
Shenoy AR, Sreenath N, Podobnik M, et al., 2005, The Rv0805 gene from Mycobacterium tuberculosis encodes a 3 ',5 '-cyclic nucleotide phosphodiesterase: Biochemical and mutational analysis, BIOCHEMISTRY, Vol: 44, Pages: 15695-15704, ISSN: 0006-2960
Shenoy AR, Srinivas A, Mahalingam M, et al., 2005, An adenylyl cyclase pseudogene in Mycobacterium tuberculosis has a functional ortholog in Mycobacterium avium, BIOCHIMIE, Vol: 87, Pages: 557-563, ISSN: 0300-9084
Shenoy AR, Sreenath NP, Mahalingam M, et al., 2005, Characterization of phylogenetically distant members of the adenylate cyclase family from mycobacteria: Rv1647 from Mycobacterium tuberculosis and its orthologue ML1399 from M. leprae, BIOCHEMICAL JOURNAL, Vol: 387, Pages: 541-551, ISSN: 0264-6021
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