Pyrin is a cytosolic pattern‐recognition receptor that normally functions as a guard to trigger capase‐1 inflammasome assembly in response to bacterial toxins and effectors that inactivate RhoA. The ...MEFV gene encoding human pyrin is preferentially expressed in phagocytes. Key domains in pyrin include a pyrin domain (PYD), a linker region, and a B30.2 domain. Binding of ASC to pyrin by a PYD‐PYD interaction triggers inflammasome assembly. Pyrin is held in an inactive conformation by negative regulation mechanisms to avoid premature inflammasome assembly. One mechanism of negative regulation involves phosphorylation of the linker by PRK kinase which in turn is positively regulated by active RhoA. The B30.2 domain also negatively regulates pyrin. Gain of function mutations in MEFV responsible for the autoinflammatory disease Familial Mediterranean Fever (FMF) map to exon 10 encoding the B30.2 domain. Insights into pyrin regulation have come from studies of several Yersinia effectors, which are injected into phagocytes and interact with the RhoA‐PRK‐pyrin axis during infection. Two effectors, YopE and YopT, inactivate RhoA to disrupt phagocytic signaling. To counteract an effector‐triggered immune response, a third effector, YopM, binds to and inhibits pyrin by hijacking PRK and RSK and directing linker phosphorylation. Inhibition of pyrin by YopM is required for virulence of Yersinia pestis, the agent of plague. Recent results from infection studies with human phagocytes and mice producing pyrin B30.2 FMF variants show that gain of function MEFV mutations bypass inhibition by YopM. Population genetic data suggest that MEFV mutations were selected for in individuals of Mediterranean decent during historic plague pandemics. This review discusses current concepts of pyrin regulation and its interaction with Yersinia effectors.
Salmonella exploit host-derived nitrate for growth in the lumen of the inflamed intestine. The generation of host-derived nitrate is dependent on Nos2, which encodes inducible nitric oxide synthase ...(iNOS), an enzyme that catalyzes nitric oxide (NO) production. However, the cellular sources of iNOS and, therefore, NO-derived nitrate used by Salmonella for growth in the lumen of the inflamed intestine remain unidentified. Here, we show that iNOS-producing inflammatory monocytes infiltrate ceca of mice infected with Salmonella. In addition, we show that inactivation of type-three secretion system (T3SS)-1 and T3SS-2 renders Salmonella unable to induce CC- chemokine receptor-2- and CC-chemokine ligand-2-dependent inflammatory monocyte recruitment. Furthermore, we show that the severity of the pathology of Salmonella- induced colitis as well as the nitrate-dependent growth of Salmonella in the lumen of the inflamed intestine are reduced in mice that lack Ccr2 and, therefore, inflammatory monocytes in the tissues. Thus, inflammatory monocytes provide a niche for Salmonella expansion in the lumen of the inflamed intestine.
Pathogenic Yersinia, including Y. pestis, the agent of plague in humans, and Y. pseudotuberculosis, the related enteric pathogen, deliver virulence effectors into host cells via a prototypical type ...III secretion system to promote pathogenesis. These effectors, termed Yersinia outer proteins (Yops), modulate multiple host signaling responses. Studies in Y. pestis and Y. pseudotuberculosis have shown that YopM suppresses infection-induced inflammasome activation; however, the underlying molecular mechanism is largely unknown. Here we show that YopM specifically restricts the pyrin inflammasome, which is triggered by the RhoA-inactivating enzymatic activities of YopE and YopT, in Y. pseudotuberculosis-infected macrophages. The attenuation of a yopM mutant is fully reversed in pyrin knockout mice, demonstrating that YopM inhibits pyrin to promote virulence. Mechanistically, YopM recruits and activates the host kinases PRK1 and PRK2 to negatively regulate pyrin by phosphorylation. These results show how a virulence factor can hijack host kinases to inhibit effector-triggered pyrin inflammasome activation.
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•Inactivation of RhoA by Yersinia effectors YopE and YopT triggers the pyrin inflammasome•Pyrin activation is blocked by YopM, an effector that binds to RSK and PRK kinases•YopM hijacks PRKs, which regulate pyrin by phosphorylation of 14-3-3 binding sites•Inhibition of the pyrin inflammasome by YopM is essential for Yersinia virulence
Pathogenic Yersinia secrete effectors called Yops, which can trigger or inhibit protective immune responses. Chung et al. demonstrate that both YopE and YopT inactivate RhoA, resulting in activation of the pyrin inflammasome. Consequently, Yersinia maintain virulence by delivering YopM, which hijacks host kinases to phosphorylate pyrin and inhibit inflammasome activation.
A) RhoA regulation of pyrin. B) Inactivation of RhoA and control of pyrin inflammasome activation by bacterial effectors and toxins.
A) In steady-state conditions, RhoA is constantly being turned on ...and off by GEFs and GAPs that exchange GDP for GTP or accelerate conversion of GTP to GDP, respectively. RhoA regulates the pyrin inflammasome through interactions with transducers of RhoA signaling, PRKs, that phosphorylate pyrin creating a 14-3-3 binding site. Phosphorylated and 14-3-3 bound pyrin is locked in an off state where its conformation prevents interaction with inflammasome components. B) (1) Bacteria deliver toxins or effecter proteins to host cells using T3SS, T6SS, secretion via ECVs or other general secretion mechanisms. There are several classes of RhoA modifying toxins including, those that covalently modify RhoA, those that proteolytically cleave RhoA and GAPs. (2) Toxins and effectors inactivate RhoA stopping PRK signaling to pyrin. (3) The lack of negative signal by PRKs and possible action of a phosphatase leads to the dephosphorylation of pyrin on its regulatory serine residues and loss of 14-3-3 binding. (4) Another class of toxins and effectors are those that inhibit pyrin inflammasome formation. YopM, an inhibitor of pyrin inflammasome formation represented in blue, recruits kinases RSK and PRK to pyrin to keep it phosphorylated. (5) When pyrin is dephosphorylated it then binds to the ASC adapter protein which binds and dimerizes pro-caspase-1 forming the inflammasome complex. Additionally, evidence suggests that microtubule polymerization is required for inflammasome formation. Upon dimerization, pro-caspase-1 undergoes autoproteolytic cleavage fully activating the enzyme. Mature caspase-1 then cleaves pro-IL-1β, pro-IL-18 and GSDMD to their mature forms. (6) Gasdermin-D then oligomerizes and forms pores in the plasma membrane which allows secretion of IL-1β and IL-18. If enough pores in the plasma membrane form the cell will undergo cell death via lysis known as pyroptosis.
Abbreviations: GDP: guanosine diphosphate; GTP: guanosine triphosphate; PRK: protein kinase C-related kinase; T3SS: type 3 secretion system; T6SS: type 6 secretion system; ECVs: extracellular vesicles; GAP: GTPase activating proteins; ASC: apoptosis-associated speck-like protein; IL: interleukin; GSDMD: gasdermin-D; PSP: protein serine/threonine phosphatase.
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•The pyrin inflammasome is triggered in response to RhoA inactivation.•Bacterial effectors and toxins that inactivate RhoA vary in mechanism.•Pyrin-induced inflammation protects against systemic bacterial infection.•Familial Mediterranean Fever is caused by mutations in the pyrin gene MEFV.•Selective pressure by plague may have driven gain of function MEFV mutations.
Pyrin is an inflammasome sensor in phagocytes that is activated in response to bacterial toxins and effectors that modify RhoA. Pathogen effector-triggered pyrin activation is analogous to an indirect guard mechanism in plants. Pyrin activation appears to be triggered when RhoA GTPases in a host cell are prevented from binding downstream signaling proteins (transducers). RhoA transducers that control this response include PRK kinases, which negatively regulate pyrin by phosphorylation and binding of 14-3-3 proteins. Microtubules regulate pyrin at different levels and may serve as a platform for inflammasome nucleation. Pyrin increases inflammation in the lung, gut or systemically during infection or intoxication in mouse models and protects against systemic infection by decreasing bacterial loads. Pathogenic Yersinia spp. overcome this protective response using effectors that inhibit the pyrin inflammasome. Gain of function mutations in MEFV, the gene encoding pyrin, cause the autoinflammatory disease Familial Mediterranean Fever. Yersinia pestis may have selected for gain of function MEFV mutations in the human population.
A type III secretion system (TTSS) is encoded on a virulence plasmid that is common to three pathogenic Yersinia species: Y. enterocolitica, Y. pseudotuberculosis, and Y. pestis. Pathogenic Yersinia ...species require this TTSS to survive and replicate within lymphoid tissues of their animal or human hosts. A set of pathogenicity factors, including those known as Yersinia outer proteins (Yops), is exported by this system upon bacterial infection of host cells. Two translocator Yops (YopB and YopD) insert into the host plasma membrane and function to transport six effector Yops (YopO, YopH, YopM, YopT, YopJ, and YopE) into the cytosol of the host cell. Effector Yops function to counteract multiple signaling responses in the infected host cell. The signaling responses counteracted by Yops are initiated by phagocytic receptors, Toll-like receptors, translocator Yops, and additional mechanisms. Innate and adaptive immune responses are thwarted as a consequence of Yop activities. A biochemical function for each effector Yop has been established, and the importance of these proteins for the pathogenesis process is being elucidated. This review focuses on the biochemical functions of Yops, the signaling pathways they modulate, and the role of these proteins in Yersinia virulence.
Yersinia outer protein J (YopJ) is a type III secretion system (T3SS) effector of pathogenic Yersinia (Yersinia pestis, Yersinia enterocolitica and Yersinia pseudotuberculosis) that is secreted into ...host cells. YopJ inhibits survival response pathways in macrophages, causing cell death. Allelic variation of YopJ is responsible for differential cytotoxicity in Yersinia strains. YopJ isoforms in Y. enterocolitica O:8 (YopP) and Y. pestis KIM (YopJ(KIM)) strains have high cytotoxic activity. In addition, YopJ(KIM)-induced macrophage death is associated with caspase-1 activation and interleukin-1β (IL-1β secretion. Here, the mechanism of YopJ(KIM)-induced cell death, caspase-1 activation, and IL-1β secretion in primary murine macrophages was examined. Caspase-3/7 activity was low and the caspase-3 substrate poly (ADP-ribose) polymerase (PARP) was not cleaved in Y. pestis KIM5-infected macrophages. In addition, cytotoxicity and IL-1β secretion were not reduced in the presence of a caspase-8 inhibitor, or in B-cell lymphoma 2 (Bcl-2)-associated X protein (Bax)/Bcl-2 homologous antagonist/killer (Bak) knockout macrophages, showing that YopJ(KIM)-mediated cell death and caspase-1 activation occur independent of mitochondrial-directed apoptosis. KIM5-infected macrophages released high mobility group protein B1 (HMGB1), a marker of necrosis, and microscopic analysis revealed that necrotic cells contained active caspase-1, indicating that caspase-1 activation is associated with necrosis. Inhibitor studies showed that receptor interacting protein 1 (RIP1) kinase and reactive oxygen species (ROS) were not required for cytotoxicity or IL-β release in KIM5-infected macrophages. IL-1β secretion was reduced in the presence of cathepsin B inhibitors, suggesting that activation of caspase-1 requires cathepsin B activity. Ectopically-expressed YopP caused higher cytotoxicity and secretion of IL-1β in Y. pseudotuberculosis-infected macrophages than YopJ(KIM). Wild-type and congenic caspase 1 knockout C57BL/6 mice were equally susceptible to lethal infection with Y. pseudotuberculosis ectopically expressing YopP. These data suggest that YopJ-induced caspase-1 activation in Yersinia-infected macrophages is a downstream consequence of necrotic cell death and is dispensable for innate host resistance to a strain with enhanced cytotoxicity.
Summary Background Yersinia pestis has caused at least three human plague pandemics. The second (Black Death, 14–17th centuries) and third (19–20th centuries) have been genetically characterised, but ...there is only a limited understanding of the first pandemic, the Plague of Justinian (6–8th centuries). To address this gap, we sequenced and analysed draft genomes of Y pestis obtained from two individuals who died in the first pandemic. Methods Teeth were removed from two individuals (known as A120 and A76) from the early medieval Aschheim-Bajuwarenring cemetery (Aschheim, Bavaria, Germany). We isolated DNA from the teeth using a modified phenol-chloroform method. We screened DNA extracts for the presence of the Y pestis -specific pla gene on the pPCP1 plasmid using primers and standards from an established assay, enriched the DNA, and then sequenced it. We reconstructed draft genomes of the infectious Y pestis strains, compared them with a database of genomes from 131 Y pestis strains from the second and third pandemics, and constructed a maximum likelihood phylogenetic tree. Findings Radiocarbon dating of both individuals (A120 to 533 AD plus or minus 98 years; A76 to 504 AD plus or minus 61 years) places them in the timeframe of the first pandemic. Our phylogeny contains a novel branch (100% bootstrap at all relevant nodes) leading to the two Justinian samples. This branch has no known contemporary representatives, and thus is either extinct or unsampled in wild rodent reservoirs. The Justinian branch is interleaved between two extant groups, 0.ANT1 and 0.ANT2, and is distant from strains associated with the second and third pandemics. Interpretation We conclude that the Y pestis lineages that caused the Plague of Justinian and the Black Death 800 years later were independent emergences from rodents into human beings. These results show that rodent species worldwide represent important reservoirs for the repeated emergence of diverse lineages of Y pestis into human populations. Funding McMaster University, Northern Arizona University, Social Sciences and Humanities Research Council of Canada, Canada Research Chairs Program, US Department of Homeland Security, US National Institutes of Health, Australian National Health and Medical Research Council.
Familial Mediterranean fever (FMF) is an autoinflammatory disease caused by homozygous or compound heterozygous gain-of-function mutations in MEFV, which encodes pyrin, an inflammasome protein. ...Heterozygous carrier frequencies for multiple MEFV mutations are high in several Mediterranean populations, suggesting that they confer selective advantage. Among 2,313 Turkish people, we found extended haplotype homozygosity flanking FMF-associated mutations, indicating evolutionarily recent positive selection of FMF-associated mutations. Two pathogenic pyrin variants independently arose >1,800 years ago. Mutant pyrin interacts less avidly with Yersinia pestis virulence factor YopM than with wild-type human pyrin, thereby attenuating YopM-induced interleukin (IL)-1β suppression. Relative to healthy controls, leukocytes from patients with FMF harboring homozygous or compound heterozygous mutations and from asymptomatic heterozygous carriers released heightened IL-1β specifically in response to Y. pestis. Y. pestis-infected Mefv
FMF knock-in mice exhibited IL-1-dependent increased survival relative to wild-type knock-in mice. Thus, FMF mutations that were positively selected in Mediterranean populations confer heightened resistance to Y. pestis.
Pathogenic
species deliver Yop effector proteins through a type III secretion system into host cells. Among these effectors, YopE and YopT are Rho-modifying toxins, which function to modulate host ...cell physiology and evade immune responses. YopE is a GTPase-activating protein (GAP) while YopT is a protease, and they inhibit RhoA by different modes of action. Modifications to RhoA are sensed by pyrin, which, once activated, assembles a caspase-1 inflammasome, which generates cytokines such as interleukin-1β (IL-1β) and cell death by pyroptosis. In
-infected macrophages, YopE or YopT triggers inflammasome assembly only in the absence of another effector, YopM, which counteracts pyrin by keeping it inactive. The glucosyltransferase TcdB from
, a well-studied RhoA-inactivating toxin, triggers activation of murine pyrin by dephosphorylation of Ser205 and Ser241. To determine if YopE or YopT triggers pyrin dephosphorylation, we infected lipopolysaccharide (LPS)-primed murine macrophages with Δ
strains expressing wild-type (wt) or YopE mutant variants or YopT. By immunoblotting pyrin after infection, we observed that wt YopE triggered dephosphorylation of Ser205 and inflammasome activation. Pyrin dephosphorylation was reduced if a YopE variant had a defect in stability or RhoA specificity but not membrane localization. We also observed that wt YopT triggered pyrin dephosphorylation but more slowly than YopE, suggesting that YopE is dominant in this process. Our findings provide evidence that RhoA-modifying toxins trigger activation of pyrin by a conserved dephosphorylation mechanism. In addition, by characterization of YopE and YopT, we show that different features of effectors, such as RhoA specificity, affect the efficiency of pyrin dephosphorylation.
White adipose tissue bridges body organs and plays a fundamental role in host metabolism. To what extent adipose tissue also contributes to immune surveillance and long-term protective defense ...remains largely unknown. Here, we have shown that at steady state, white adipose tissue contained abundant memory lymphocyte populations. After infection, white adipose tissue accumulated large numbers of pathogen-specific memory T cells, including tissue-resident cells. Memory T cells in white adipose tissue expressed a distinct metabolic profile, and white adipose tissue from previously infected mice was sufficient to protect uninfected mice from lethal pathogen challenge. Induction of recall responses within white adipose tissue was associated with the collapse of lipid metabolism in favor of antimicrobial responses. Our results suggest that white adipose tissue represents a memory T cell reservoir that provides potent and rapid effector memory responses, positioning this compartment as a potential major contributor to immunological memory.
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•White adipose tissue serves as a reservoir for memory T cells•White adipose tissue memory T cells have a distinct functional and metabolic profile•Memory T cells from white adipose tissue can protect against lethal infectious challenge•Reactivation of white adipose tissue memory T cells alters adipose tissue physiology
The role of adipose tissue in protective immunity is largely unknown. Han et al. reveal that white adipose tissue is a reservoir for memory T cells endowed with a distinct functional and metabolic profile. These memory T cells are able to protect against infection while inducing physiological remodeling of adipose tissue.
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