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Ashley Mansell - One of the best experts on this subject based on the ideXlab platform.
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myd88 adapter like mal Tirap interaction with traf6 is critical for tlr2 and tlr4 mediated nf kappab proinflammatory responses
Journal of Biological Chemistry, 2009Co-Authors: Brett Verstak, Kamalpreet Nagpal, Stephen P Bottomley, Douglas T Golenbock, Paul J Hertzog, Ashley MansellAbstract:Toll/interleukin-1 (Tir)Receptor-containing adapters are critical in orchestrating the different signal transduction pathways following Toll-like Receptor (TLR) activation. MyD88 adapter-like (Mal), also termed TirAP, is involved in bridging MyD88 to the Receptor complex for TLR-2 and TLR4 signaling in response to bacterial infection. We have previously reported an interaction between Mal and tumor necrosis factor Receptor-associated factor 6 (TRAF6) via a TRAF6-binding motif, the disruption of which inhibited TLR-mediated NF-kappaB-luciferase reporter activity. Given the recent report of intracellular TRAM localization promoting sequential signaling in TLR4 responses, we further characterized Mal interaction with TRAF6, the cellular localization, and the outcomes of disrupting this association on TLR inflammatory responses. We found that Mal and TRAF6 directly interact in response to TLR2 and TLR4 stimulation, although membrane localization is not necessary to facilitate interaction. Critically, reconstitution of murine Mal-deficient macrophages with MalE190A, containing a mutation within the TRAF6-binding motif, fails to reconstitute the proinflammatory response to TLR2 and TLR4 ligands compared with wild type Mal. Furthermore, Mal interaction with TRAF6 mediates Ser phosphorylation of the p65 subunit of NF-kappaB and thus controls transcriptional activation but not nuclear translocation of NF-kappaB. This study characterizes the novel role for Mal in facilitating the direct recruitment of TRAF6 to the plasma membrane, which is necessary for TLR2- and TLR4-induced transactivation of NF-kappaB and regulation of the subsequent pro-inflammatory response.
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myd88 adapter like mal Tirap interaction with traf6 is critical for tlr2 and tlr4 mediated nf κb proinflammatory responses
Journal of Biological Chemistry, 2009Co-Authors: Brett Verstak, Kamalpreet Nagpal, Stephen P Bottomley, Douglas T Golenbock, Paul J Hertzog, Ashley MansellAbstract:Toll/interleukin-1 (Tir)Receptor-containing adapters are critical in orchestrating the different signal transduction pathways following Toll-like Receptor (TLR) activation. MyD88 adapter-like (Mal), also termed TirAP, is involved in bridging MyD88 to the Receptor complex for TLR-2 and TLR4 signaling in response to bacterial infection. We have previously reported an interaction between Mal and tumor necrosis factor Receptor-associated factor 6 (TRAF6) via a TRAF6-binding motif, the disruption of which inhibited TLR-mediated NF-κB-luciferase reporter activity. Given the recent report of intracellular TRAM localization promoting sequential signaling in TLR4 responses, we further characterized Mal interaction with TRAF6, the cellular localization, and the outcomes of disrupting this association on TLR inflammatory responses. We found that Mal and TRAF6 directly interact in response to TLR2 and TLR4 stimulation, although membrane localization is not necessary to facilitate interaction. Critically, reconstitution of murine Mal-deficient macrophages with MalE190A, containing a mutation within the TRAF6-binding motif, fails to reconstitute the proinflammatory response to TLR2 and TLR4 ligands compared with wild type Mal. Furthermore, Mal interaction with TRAF6 mediates Ser phosphorylation of the p65 subunit of NF-κB and thus controls transcriptional activation but not nuclear translocation of NF-κB. This study characterizes the novel role for Mal in facilitating the direct recruitment of TRAF6 to the plasma membrane, which is necessary for TLR2- and TLR4-induced transactivation of NF-κB and regulation of the subsequent pro-inflammatory response.
Brendan Kenny - One of the best experts on this subject based on the ideXlab platform.
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enteropathogenic escherichia coli epec Tir Receptor molecule does not undergo full modification when introduced into host cells by epec independent mechanisms
Infection and Immunity, 2001Co-Authors: Brendan Kenny, Jonathan WarawaAbstract:Enteropathogenic Escherichia coli (EPEC), like many other gram-negative pathogens, encodes a type III secretion apparatus dedicated to the release of virulence-associated proteins. One such protein, Tir, is translocated into host cells, where it is modified by the addition of phosphate groups, resulting in a number of species with distinct molecular mass. One phosphorylation event, on tyrosine residue 474 of Tir, does not contribute to shifts in molecular mass but is essential for its actin-nucleating function. The role of the nonphosphotyrosine related modifications is unknown. In this paper, we demonstrate, using three different approaches, that Tir does not encode sufficient information to facilitate its complete modification when introduced into host cells in EPEC-independent mechanisms. Each system revealed that Tir is a substrate for a host kinase whose action results in its partial modification to a form similar to one evident in EPEC-infected host cells. Further Tir modification could not be induced by infecting cells with EPEC, suggesting that Tir must be coexpressed with other EPEC factors to enable its full modification within host cells. One approach used Yersinia spp. to deliver Tir into host cells, and this system revealed that Tir secretion and translocation can occur in the absence of the Tir chaperone molecule, CesT (formerly known as OrfU). CesT was found to be an efficiency factor which was not required, unlike in EPEC, for Tir stability, indicating that it may function to guide Tir to the translocation apparatus or maintain it in a secretion-competent form.
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phosphorylation of tyrosine 474 of the enteropathogenic escherichia coli epec Tir Receptor molecule is essential for actin nucleating activity and is preceded by additional host modifications
Molecular Microbiology, 1999Co-Authors: Brendan KennyAbstract:The enteropathogenic Escherichia coli (EPEC) Tir protein becomes tyrosine phosphorylated in host cells and displays an increase in apparent molecular mass. The interaction of Tir with the EPEC outer membrane protein, intimin, triggers actin nucleation beneath the adherent bacteria. The enterohaemorrhagic E. coli O157:H7 (EHEC) Tir molecule is not tyrosine phosphorylated. In this paper, Tir tyrosine phosphorylation is shown to be essential for actin nucleation activity, but not for the increase in apparent molecular mass observed in target cells. Tyrosine phosphorylation had no role in Tir molecular mass shift, indicating additional host modifications. Analysis of Tir intermediates indicates that tyrosine-independent modification functions to direct Tir's correct insertion from the cytoplasm into the host membrane. Deletion analysis identified Tir domains participating in translocation, association with the host membrane, modification and antibody recognition. Intimin was found to bind a 55-amino-acid region (TIBA) within Tir that topological and sequence analysis suggests is located in an extracellular loop. Homologous TIBA sequences exist in integrins, which also bind intimin. Collectively, this study provides definitive evidence for the importance of tyrosine phosphorylation for EPEC Tir function and reveals differences in the pathogenicity of EPEC and EHEC. The data also suggest a mechanism for Tir insertion into the host membrane, as well as providing clues to the mode of intimin-integrin interaction.
Brett Verstak - One of the best experts on this subject based on the ideXlab platform.
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myd88 adapter like mal Tirap interaction with traf6 is critical for tlr2 and tlr4 mediated nf kappab proinflammatory responses
Journal of Biological Chemistry, 2009Co-Authors: Brett Verstak, Kamalpreet Nagpal, Stephen P Bottomley, Douglas T Golenbock, Paul J Hertzog, Ashley MansellAbstract:Toll/interleukin-1 (Tir)Receptor-containing adapters are critical in orchestrating the different signal transduction pathways following Toll-like Receptor (TLR) activation. MyD88 adapter-like (Mal), also termed TirAP, is involved in bridging MyD88 to the Receptor complex for TLR-2 and TLR4 signaling in response to bacterial infection. We have previously reported an interaction between Mal and tumor necrosis factor Receptor-associated factor 6 (TRAF6) via a TRAF6-binding motif, the disruption of which inhibited TLR-mediated NF-kappaB-luciferase reporter activity. Given the recent report of intracellular TRAM localization promoting sequential signaling in TLR4 responses, we further characterized Mal interaction with TRAF6, the cellular localization, and the outcomes of disrupting this association on TLR inflammatory responses. We found that Mal and TRAF6 directly interact in response to TLR2 and TLR4 stimulation, although membrane localization is not necessary to facilitate interaction. Critically, reconstitution of murine Mal-deficient macrophages with MalE190A, containing a mutation within the TRAF6-binding motif, fails to reconstitute the proinflammatory response to TLR2 and TLR4 ligands compared with wild type Mal. Furthermore, Mal interaction with TRAF6 mediates Ser phosphorylation of the p65 subunit of NF-kappaB and thus controls transcriptional activation but not nuclear translocation of NF-kappaB. This study characterizes the novel role for Mal in facilitating the direct recruitment of TRAF6 to the plasma membrane, which is necessary for TLR2- and TLR4-induced transactivation of NF-kappaB and regulation of the subsequent pro-inflammatory response.
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myd88 adapter like mal Tirap interaction with traf6 is critical for tlr2 and tlr4 mediated nf κb proinflammatory responses
Journal of Biological Chemistry, 2009Co-Authors: Brett Verstak, Kamalpreet Nagpal, Stephen P Bottomley, Douglas T Golenbock, Paul J Hertzog, Ashley MansellAbstract:Toll/interleukin-1 (Tir)Receptor-containing adapters are critical in orchestrating the different signal transduction pathways following Toll-like Receptor (TLR) activation. MyD88 adapter-like (Mal), also termed TirAP, is involved in bridging MyD88 to the Receptor complex for TLR-2 and TLR4 signaling in response to bacterial infection. We have previously reported an interaction between Mal and tumor necrosis factor Receptor-associated factor 6 (TRAF6) via a TRAF6-binding motif, the disruption of which inhibited TLR-mediated NF-κB-luciferase reporter activity. Given the recent report of intracellular TRAM localization promoting sequential signaling in TLR4 responses, we further characterized Mal interaction with TRAF6, the cellular localization, and the outcomes of disrupting this association on TLR inflammatory responses. We found that Mal and TRAF6 directly interact in response to TLR2 and TLR4 stimulation, although membrane localization is not necessary to facilitate interaction. Critically, reconstitution of murine Mal-deficient macrophages with MalE190A, containing a mutation within the TRAF6-binding motif, fails to reconstitute the proinflammatory response to TLR2 and TLR4 ligands compared with wild type Mal. Furthermore, Mal interaction with TRAF6 mediates Ser phosphorylation of the p65 subunit of NF-κB and thus controls transcriptional activation but not nuclear translocation of NF-κB. This study characterizes the novel role for Mal in facilitating the direct recruitment of TRAF6 to the plasma membrane, which is necessary for TLR2- and TLR4-induced transactivation of NF-κB and regulation of the subsequent pro-inflammatory response.
Julia Weikum - One of the best experts on this subject based on the ideXlab platform.
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the extracellular juncture domains in the intimin passenger adopt a constitutively extended conformation inducing restraints to its sphere of action
Scientific Reports, 2020Co-Authors: Monika Schütz, Julia Weikum, Alina Kulakova, Giulio Tesei, Shogo Yoshimoto, Line Vejby Jaegerum, Katsutoshi Hori, Marie Skepo, Pernille HarrisAbstract:Enterohemorrhagic and enteropathogenic Escherichia coli are among the most important food-borne pathogens, posing a global health threat. The virulence factor intimin is essential for the attachment of pathogenic E. coli to the intestinal host cell. Intimin consists of four extracellular bacterial immunoglobulin-like (Big) domains, D00–D2, extending into the fifth lectin subdomain (D3) that binds to the Tir-Receptor on the host cell. Here, we present the crystal structures of the elusive D00–D0 domains at 1.5 A and D0–D1 at 1.8 A resolution, which confirms that the passenger of intimin has five distinct domains. We describe that D00–D0 exhibits a higher degree of rigidity and D00 likely functions as a juncture domain at the outer membrane-extracellular medium interface. We conclude that D00 is a unique Big domain with a specific topology likely found in a broad range of other inverse autotransporters. The accumulated data allows us to model the complete passenger of intimin and propose functionality to the Big domains, D00–D0–D1, extending directly from the membrane.
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the extracellular juncture domains in the intimin passenger adopt a constitutively extended conformation inducing restraints to its sphere of action
Social Science Research Network, 2020Co-Authors: Julia Weikum, Monika Schütz, Alina Kulakova, Giulio Tesei, Shogo Yoshimoto, Line Vejby Jaegerum, Katsutoshi Hori, Marie Skepo, Pernille Harris, Jack C. LeoAbstract:Enterohemorrhagic and enteropathogenic Escherichia coli are among the most important food-borne pathogens, posing a global health threat. The virulence factor intimin is essential for attachment of pathogenic E. coli to the intestinal host cell. Intimin consists of four extracellular bacterial immunoglobulin-like (Big) domains, D00-D2, extending into the fifth lectin subdomain (D3) that binds to the Tir-Receptor on the host cell. Here, we present the crystal structures of the elusive D00-D0 domains at 1.5 A and D0-D1 at 1.8 A resolution that confirm that the passenger of intimin has five distinct domains. We describe that D00-D0 exhibits a higher degree of rigidity and D00 likely functions as a juncture domain at the outer membrane-extracellular medium interface. We conclude that D00 is a unique Big domain with a specific topology likely found in a broad range of other inverse autotransporters. The accumulated data allows us to model the complete passenger of intimin and propose functionality to the Big domains, D00-D0-D1, extending directly from the membrane.
Kamalpreet Nagpal - One of the best experts on this subject based on the ideXlab platform.
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myd88 adapter like mal Tirap interaction with traf6 is critical for tlr2 and tlr4 mediated nf kappab proinflammatory responses
Journal of Biological Chemistry, 2009Co-Authors: Brett Verstak, Kamalpreet Nagpal, Stephen P Bottomley, Douglas T Golenbock, Paul J Hertzog, Ashley MansellAbstract:Toll/interleukin-1 (Tir)Receptor-containing adapters are critical in orchestrating the different signal transduction pathways following Toll-like Receptor (TLR) activation. MyD88 adapter-like (Mal), also termed TirAP, is involved in bridging MyD88 to the Receptor complex for TLR-2 and TLR4 signaling in response to bacterial infection. We have previously reported an interaction between Mal and tumor necrosis factor Receptor-associated factor 6 (TRAF6) via a TRAF6-binding motif, the disruption of which inhibited TLR-mediated NF-kappaB-luciferase reporter activity. Given the recent report of intracellular TRAM localization promoting sequential signaling in TLR4 responses, we further characterized Mal interaction with TRAF6, the cellular localization, and the outcomes of disrupting this association on TLR inflammatory responses. We found that Mal and TRAF6 directly interact in response to TLR2 and TLR4 stimulation, although membrane localization is not necessary to facilitate interaction. Critically, reconstitution of murine Mal-deficient macrophages with MalE190A, containing a mutation within the TRAF6-binding motif, fails to reconstitute the proinflammatory response to TLR2 and TLR4 ligands compared with wild type Mal. Furthermore, Mal interaction with TRAF6 mediates Ser phosphorylation of the p65 subunit of NF-kappaB and thus controls transcriptional activation but not nuclear translocation of NF-kappaB. This study characterizes the novel role for Mal in facilitating the direct recruitment of TRAF6 to the plasma membrane, which is necessary for TLR2- and TLR4-induced transactivation of NF-kappaB and regulation of the subsequent pro-inflammatory response.
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myd88 adapter like mal Tirap interaction with traf6 is critical for tlr2 and tlr4 mediated nf κb proinflammatory responses
Journal of Biological Chemistry, 2009Co-Authors: Brett Verstak, Kamalpreet Nagpal, Stephen P Bottomley, Douglas T Golenbock, Paul J Hertzog, Ashley MansellAbstract:Toll/interleukin-1 (Tir)Receptor-containing adapters are critical in orchestrating the different signal transduction pathways following Toll-like Receptor (TLR) activation. MyD88 adapter-like (Mal), also termed TirAP, is involved in bridging MyD88 to the Receptor complex for TLR-2 and TLR4 signaling in response to bacterial infection. We have previously reported an interaction between Mal and tumor necrosis factor Receptor-associated factor 6 (TRAF6) via a TRAF6-binding motif, the disruption of which inhibited TLR-mediated NF-κB-luciferase reporter activity. Given the recent report of intracellular TRAM localization promoting sequential signaling in TLR4 responses, we further characterized Mal interaction with TRAF6, the cellular localization, and the outcomes of disrupting this association on TLR inflammatory responses. We found that Mal and TRAF6 directly interact in response to TLR2 and TLR4 stimulation, although membrane localization is not necessary to facilitate interaction. Critically, reconstitution of murine Mal-deficient macrophages with MalE190A, containing a mutation within the TRAF6-binding motif, fails to reconstitute the proinflammatory response to TLR2 and TLR4 ligands compared with wild type Mal. Furthermore, Mal interaction with TRAF6 mediates Ser phosphorylation of the p65 subunit of NF-κB and thus controls transcriptional activation but not nuclear translocation of NF-κB. This study characterizes the novel role for Mal in facilitating the direct recruitment of TRAF6 to the plasma membrane, which is necessary for TLR2- and TLR4-induced transactivation of NF-κB and regulation of the subsequent pro-inflammatory response.