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Swacha, P., Gekara, N. O. & Erttmann, S. F. (2019). Biochemical and microscopic analysis of inflammasome complex formation. In: Jungsan Sohn (Ed.), DNA Sensors and Inflammasomes: (pp. 287-298). Elsevier, 625, Article ID S0076-6879(19)30173-9.
Öppna denna publikation i ny flik eller fönster >>Biochemical and microscopic analysis of inflammasome complex formation
2019 (Engelska)Ingår i: DNA Sensors and Inflammasomes / [ed] Jungsan Sohn, Elsevier, 2019, Vol. 625, s. 287-298, artikel-id S0076-6879(19)30173-9Kapitel i bok, del av antologi (Refereegranskat)
Abstract [en]

Inflammasomes are multiprotein signaling platforms responsible for the maturation of pro-IL-1β and pro-IL-18 as well as the induction of an inflammatory cell death termed pyroptosis. Most inflammasomes consist of an upstream sensor, in most cases an adaptor protein (ASC) and inflammatory caspases such as caspase-1. Upon activation, sensor proteins oligomerize with adaptor proteins, forming large complexes called specks. These complexes can be stabilized and detected by Western blotting or fluorescence microscopy providing a direct evidence of inflammasome activation. Here we describe protocols for two complementary methods for detecting inflammasome complexes: (1) biochemical isolation and detection of ASC oligomers by Western blot analysis and (2) microscopic visualization of active caspase-1—ASC complexes. These protocols have successfully been applied in our recent study to unveil new regulatory mechanisms for different inflammasomes including the DNA sensor AIM2 (Erttmann et al., 2016).

Ort, förlag, år, upplaga, sidor
Elsevier, 2019
Serie
Methods in Enzymology, ISSN 0076-6879 ; 625
Nyckelord
Inflammasome complex, ASC specks, Caspase 1, AIM2, IL-1β, Innate immunity
Nationell ämneskategori
Cell- och molekylärbiologi
Forskningsämne
immunologi; cellforskning; infektionssjukdomar
Identifikatorer
urn:nbn:se:umu:diva-162825 (URN)10.1016/bs.mie.2019.05.014 (DOI)000488782900018 ()31455532 (PubMedID)2-s2.0-85067031234 (Scopus ID)978-0-12-818359-5 (ISBN)
Tillgänglig från: 2019-08-30 Skapad: 2019-08-30 Senast uppdaterad: 2019-10-25Bibliografiskt granskad
Erttmann, S. F. & Gekara, N. O. (2019). Hydrogen peroxide release by bacteria suppresses inflammasome-dependent innate immunity. Nature Communications, 10, Article ID 3493.
Öppna denna publikation i ny flik eller fönster >>Hydrogen peroxide release by bacteria suppresses inflammasome-dependent innate immunity
2019 (Engelska)Ingår i: Nature Communications, ISSN 2041-1723, E-ISSN 2041-1723, Vol. 10, artikel-id 3493Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

Hydrogen peroxide (H2O2) has a major function in host-microbial interactions. Although most studies have focused on the endogenous H2O2 produced by immune cells to kill microbes, bacteria can also produce H2O2. How microbial H2O2 influences the dynamics of host-microbial interactions is unclear. Here we show that H2O2 released by Streptococcus pneumoniae inhibits inflammasomes, key components of the innate immune system, contributing to the pathogen colonization of the host. We also show that the oral commensal H2O2-producing bacteria Streptococcus oralis can block inflammasome activation. This study uncovers an unexpected role of H2O2 in immune suppression and demonstrates how, through this mechanism, bacteria might restrain the immune system to co-exist with the host.

Ort, förlag, år, upplaga, sidor
Nature Publishing Group, 2019
Nyckelord
Inflammasome
Nationell ämneskategori
Cell- och molekylärbiologi
Forskningsämne
immunologi; cellforskning; infektionssjukdomar
Identifikatorer
urn:nbn:se:umu:diva-162589 (URN)10.1038/s41467-019-11169-x (DOI)000478576800011 ()31375698 (PubMedID)
Tillgänglig från: 2019-08-23 Skapad: 2019-08-23 Senast uppdaterad: 2019-08-30Bibliografiskt granskad
Bamyaci, S., Ekestubbe, S., Nordfelth, R., Erttmann, S. F., Edgren, T. & Forsberg, Å. (2018). YopN Is Required for Efficient Effector Translocation and Virulence in Yersinia pseudotuberculosis. Infection and Immunity, 86(8), Article ID e00957-17.
Öppna denna publikation i ny flik eller fönster >>YopN Is Required for Efficient Effector Translocation and Virulence in Yersinia pseudotuberculosis
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2018 (Engelska)Ingår i: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 86, nr 8, artikel-id e00957-17Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

Type III secretion systems (T3SSs) are used by various Gram-negative pathogens to subvert the host defense by a host cell contact-dependent mechanism to secrete and translocate virulence effectors. While the effectors differ between pathogens and determine the pathogenic life style, the overall mechanism of secretion and translocation is conserved. T3SSs are regulated at multiple levels, and some secreted substrates have also been shown to function in regulation. In Yersinia, one of the substrates, YopN, has long been known to function in the host cell contact-dependent regulation of the T3SS. Prior to contact, through its interaction with TyeA, YopN blocks secretion. Upon cell contact, TyeA dissociates from YopN, which is secreted by the T3SS, resulting in the induction of the system. YopN has also been shown to be translocated into target cells by a T3SS-dependent mechanism. However, no intracellular function has yet been assigned to YopN. The regulatory role of YopN involves the N-terminal and C-terminal parts, while less is known about the role of the central region of YopN. Here, we constructed different in-frame deletion mutants within the central region. The deletion of amino acids 76 to 181 resulted in an unaltered regulation of Yop expression and secretion but triggered reduced YopE and YopH translocation within the first 30 min after infection. As a consequence, this deletion mutant lost its ability to block phagocytosis by macrophages. In conclusion, we were able to differentiate the function of YopN in translocation and virulence from its function in regulation.

Nyckelord
phagocytosis, type III secretion, Yersinia, YopN, virulence
Nationell ämneskategori
Mikrobiologi inom det medicinska området
Identifikatorer
urn:nbn:se:umu:diva-150354 (URN)10.1128/IAI.00957-17 (DOI)000439474900006 ()29760214 (PubMedID)
Tillgänglig från: 2018-08-13 Skapad: 2018-08-13 Senast uppdaterad: 2019-01-09Bibliografiskt granskad
Wang, H., Avican, K., Fahlgren, A., Erttmann, S. F., Nuss, A. M., Dersch, P., . . . Wolf-Watz, H. (2016). Increased plasmid copy number is essential for Yersinia T3SS function and virulence. Science, 353(6298), 492-495
Öppna denna publikation i ny flik eller fönster >>Increased plasmid copy number is essential for Yersinia T3SS function and virulence
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2016 (Engelska)Ingår i: Science, ISSN 0036-8075, E-ISSN 1095-9203, Vol. 353, nr 6298, s. 492-495Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

Pathogenic bacteria have evolved numerous virulence mechanisms that are essential for establishing infections. The enterobacterium Yersinia uses a type III secretion system (T3SS) encoded by a 70-kilobase, low-copy, IncFII-class virulence plasmid. We report a novel virulence strategy in Y. pseudotuberculosis in which this pathogen up-regulates the plasmid copy number during infection. We found that an increased dose of plasmid-encoded genes is indispensable for virulence and substantially elevates the expression and function of the T3SS. Remarkably, we observed direct, tight coupling between plasmid replication and T3SS function. This regulatory pathway provides a framework for further exploration of the environmental sensing mechanisms of pathogenic bacteria.

Nationell ämneskategori
Mikrobiologi inom det medicinska området Infektionsmedicin
Identifikatorer
urn:nbn:se:umu:diva-125586 (URN)10.1126/science.aaf7501 (DOI)000380583600042 ()27365311 (PubMedID)
Tillgänglig från: 2016-09-19 Skapad: 2016-09-13 Senast uppdaterad: 2018-06-07Bibliografiskt granskad
Erttmann, S. F., Härtlova, A., Sloniecka, M., Raffi, F. A. M., Hosseinzadeh, A., Edgren, T., . . . Gekara, N. O. (2016). Loss of the DNA Damage Repair Kinase ATM Impairs Inflammasome-Dependent Anti-Bacterial Innate Immunity. Immunity, 45(1), 106-118
Öppna denna publikation i ny flik eller fönster >>Loss of the DNA Damage Repair Kinase ATM Impairs Inflammasome-Dependent Anti-Bacterial Innate Immunity
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2016 (Engelska)Ingår i: Immunity, ISSN 1074-7613, E-ISSN 1097-4180, Vol. 45, nr 1, s. 106-118Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

The ATM kinase is a central component of the DNA damage repair machinery and redox balance. ATM dysfunction results in the multisystem disease ataxia-telangiectasia (AT). A major cause of mortality in AT is respiratory bacterial infections. Whether ATM deficiency causes innate immune defects that might contribute to bacterial infections is not known. Here we have shown that loss of ATM impairs inflammasome- dependent anti-bacterial innate immunity. Cells from AT patients or Atm(-/-) mice exhibited diminished interleukin-1 beta (IL-1 beta) production in response to bacteria. In vivo, Atm(-/-) mice were more susceptible to pulmonary S. pneumoniae infection in a manner consistent with inflammasome defects. Our data indicate that such defects were due to oxidative inhibition of inflammasome complex assembly. This study reveals an unanticipated function of reactive oxygen species (ROS) in negative regulation of inflammasomes and proposes a theory for the notable susceptibility of AT patients to pulmonary bacterial infection.

Nationell ämneskategori
Immunologi inom det medicinska området
Identifikatorer
urn:nbn:se:umu:diva-125590 (URN)10.1016/j.immuni.2016.06.018 (DOI)000380749000014 ()27421701 (PubMedID)
Tillgänglig från: 2016-09-23 Skapad: 2016-09-13 Senast uppdaterad: 2018-06-07Bibliografiskt granskad
Härtlova, A., Erttmann, S. F., Raffi, F. A. M., Schmalz, A. M., Resch, U., Anugula, S., . . . Gekara, N. O. (2015). DNA Damage Primes the Type I Interferon System via the Cytosolic DNA Sensor STING to Promote Anti-Microbial Innate Immunity. Immunity, 42(2), 332-343
Öppna denna publikation i ny flik eller fönster >>DNA Damage Primes the Type I Interferon System via the Cytosolic DNA Sensor STING to Promote Anti-Microbial Innate Immunity
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2015 (Engelska)Ingår i: Immunity, ISSN 1074-7613, E-ISSN 1097-4180, Vol. 42, nr 2, s. 332-343Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

Dysfunction in Ataxia-telangiectasia mutated (ATM), a central component of the DNA repair machinery, results in Ataxia Telangiectasia (AT), a cancer-prone disease with a variety of inflammatory manifestations. By analyzing AT patient samples and Atm(-/-) mice, we found that unrepaired DNA lesions induce type I interferons (IFNs), resulting in enhanced anti-viral and anti-bacterial responses in Atm(-/-) mice. Priming of the type I interferon system by DNA damage involved release of DNA into the cytoplasm where it activated the cytosolic DNA sensing STING-mediated pathway, which in turn enhanced responses to innate stimuli by activating the expression of Toll-like receptors, RIG-I-like receptors, cytoplasmic DNA sensors, and their downstream signaling partners. This study provides a potential explanation for the inflammatory phenotype of AT patients and establishes damaged DNA as a cell intrinsic danger signal that primes the innate immune system for a rapid and amplified response to microbial and environmental threats.

Nationell ämneskategori
Medicinsk bioteknologi (med inriktning mot cellbiologi (inklusive stamcellsbiologi), molekylärbiologi, mikrobiologi, biokemi eller biofarmaci)
Identifikatorer
urn:nbn:se:umu:diva-101396 (URN)10.1016/j.immuni.2015.01.012 (DOI)000349916400016 ()25692705 (PubMedID)
Tillgänglig från: 2015-07-02 Skapad: 2015-03-30 Senast uppdaterad: 2018-06-07Bibliografiskt granskad
Erttmann, S. F., Gekara, N. O. & Fällman, M. (2014). Bacteria Induce Prolonged PMN Survival via a Phosphatidylcholine-Specific Phospholipase C- and Protein Kinase C-Dependent Mechanism. PLoS ONE, 9(1), e87859
Öppna denna publikation i ny flik eller fönster >>Bacteria Induce Prolonged PMN Survival via a Phosphatidylcholine-Specific Phospholipase C- and Protein Kinase C-Dependent Mechanism
2014 (Engelska)Ingår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 9, nr 1, s. e87859-Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

Polymorphonuclear leukocytes (PMNs) are essential for the human innate immune defense, limiting expansion of invading microorganisms. PMN turnover is controlled by apoptosis, but the regulating signaling pathways remain elusive, largely due to inherent differences between mice and humans that undermine use of mouse models for understanding human PMN biology. Here, we aim to elucidate signal transduction mediating survival of human peripheral blood PMNs in response to bacteria, such as Yersinia pseudotuberculosis, an enteropathogen that causes the gastro-intestinal disease yersiniosis, as well as Escherichia coli and Staphylococcus aureus. Determinations of cell death reveal that uninfected control cells undergo apoptosis, while PMNs infected with either Gram-positive or -negative bacteria show profoundly increased survival. Infected cells exhibit decreased caspase 3 and 8 activities, increased mitochondrial integrity and are resistant to apoptosis induced by a death receptor ligand. This bacteria-induced response is accompanied by pro-inflammatory cytokine production including interleukin-8 and tumor necrosis factor-a competent to attract additional PMNs. Using agonists and pharmacological inhibitors, we show participation of Toll-like receptor 2 and 4, and interestingly, that protein kinase C (PKC) and phosphatidylcholine-specific phospholipase C (PC-PLC), but not tyrosine kinases or phosphatidylinositol-specific phospholipase C (PI-PLC) are key players in this dual PMN response. Our findings indicate the importance of prolonged PMN survival in response to bacteria, where general signaling pathways ensure complete exploitation of PMN anti-microbial capacity.

Nationell ämneskategori
Medicin och hälsovetenskap
Identifikatorer
urn:nbn:se:umu:diva-87041 (URN)10.1371/journal.pone.0087859 (DOI)000330621900186 ()
Tillgänglig från: 2014-04-07 Skapad: 2014-03-18 Senast uppdaterad: 2018-06-08Bibliografiskt granskad
Thorslund, S. E., Ermert, D., Fahlgren, A., Erttmann, S. F., Nilsson, K., Hosseinzadeh, A., . . . Fällman, M. (2013). Role of YopK in Yersinia pseudotuberculosis Resistance Against Polymorphonuclear Leukocyte Defense. Infection and Immunity, 81(1), 11-22
Öppna denna publikation i ny flik eller fönster >>Role of YopK in Yersinia pseudotuberculosis Resistance Against Polymorphonuclear Leukocyte Defense
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2013 (Engelska)Ingår i: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 81, nr 1, s. 11-22Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

The enteropathogen Y. pseudotuberculosis can survive in the harsh environment of lymphoid compartments that abounds in immune cells. This capacity is dependent on the plasmid-encoded Yersinia outer proteins (Yops) that are delivered into the host cell via a mechanism involving the Yersinia type three secretion system. We show that the virulence protein YopK has a role in the mechanism by which Y. pseudotuberculosis avoids the polymorphonuclear leukocyte (PMN, or neutrophil) defense. A yopK mutant, which is attenuated in the mouse infection model where it fails to cause systemic infection, was found to colonize Peyer's patches and mesenteric lymph nodes more rapidly than the wild-type strain. Further, in mice lacking PMNs, the yopK mutant caused full disease with systemic spread and typical symptoms. Analyses of effects on PMNs revealed that both the wild-type strain and the yopK mutant inhibited internalization and ROS production, as well as neutrophil extracellular trap formation by PMNs. However, the wild-type strain effectively avoided induction PMN death, whereas the mutant caused a necrotic-like PMN death. Taken together, our results indicate that YopK is required for the ability of Yersinia to resist the PMN defense, which is critical for the virulence of the pathogen. We suggest a mechanism where YopK functions to prevent unintended Yop delivery and thereby PMN disruption resulting in necrotic like cell death, which would enhance the inflammatory response favoring the host.

Nationell ämneskategori
Mikrobiologi
Identifikatorer
urn:nbn:se:umu:diva-61273 (URN)10.1128/IAI.00650-12 (DOI)000316298000002 ()23090955 (PubMedID)
Tillgänglig från: 2012-11-07 Skapad: 2012-11-07 Senast uppdaterad: 2018-06-08Bibliografiskt granskad
Bamyaci, S., Ekestubbe, S., Nordfelth, R., Ertmann, S., Edgren, T. & Forsberg, Å.YopN is required for efficient translocation and virulence in Yersinia pseudotuberculosis.
Öppna denna publikation i ny flik eller fönster >>YopN is required for efficient translocation and virulence in Yersinia pseudotuberculosis
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(Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
Nationell ämneskategori
Cell- och molekylärbiologi
Identifikatorer
urn:nbn:se:umu:diva-130153 (URN)
Tillgänglig från: 2017-01-12 Skapad: 2017-01-12 Senast uppdaterad: 2018-06-09
Organisationer
Identifikatorer
ORCID-id: ORCID iD iconorcid.org/0000-0003-2398-8405

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