scholarly journals NOD1 is super-activated through spatially-selective ubiquitination by the Salmonella effector SspH2

2021 ◽  
Author(s):  
Cole Delyea ◽  
Shu Luo ◽  
Bradley E Dubrule ◽  
Olivier Julien ◽  
Amit P Bhavsar
Keyword(s):  
Mass Spectrometry ◽  
Ubiquitin Ligase ◽  
Inflammatory Cytokine ◽  
E3 Ubiquitin Ligase ◽  
Cytokine Secretion ◽  
Host Cells ◽  
List Type ◽  
The Novel ◽  
Inflammatory Signaling ◽  
Lysine Residues

As part of its pathogenesis, Salmonella enterica serovar Typhimurium delivers effector proteins into host cells. One effector is SspH2, a member of the novel E3 ubiquitin ligase family, interacts with, and enhances, NOD1 pro-inflammatory signaling, though the underlying mechanisms are unclear. Here, we report the novel discovery that SspH2 interacts with multiple members of the NLRC family to enhance pro-inflammatory signaling that results from targeted ubiquitination. We show that SspH2 modulates host innate immunity by interacting with both NOD1 and NOD2 in mammalian epithelial cell culture. We also show that SspH2 specifically interacts with the NBD and LRR domains of NOD1 and super-activates NOD1- and NOD2-mediated cytokine secretion via the NF-κB pathway. Mass spectrometry analyses identified lysine residues in NOD1 that were ubiquitinated after interaction with SspH2. Through NOD1 mutational analyses, we identified four key lysine residues that are required for NOD1 super-activation by SspH2, but not its basal activity. These critical lysine residues are positioned in the same region of NOD1 and define a surface on NOD1 that is targeted by SspH2. Overall, this work provides evidence for post-translational modification of NOD1 by ubiquitin, and uncovers a unique mechanism of spatially-selective ubiquitination to enhance the activation of an archetypal NLR.

scholarly journals Lysine11-Linked Polyubiquitination of the AnkB F-Box Effector of Legionella pneumophila

2015 ◽  
Vol 84 (1) ◽  
pp. 99-107 ◽  
Author(s):  
William M. Bruckert ◽  
Yousef Abu Kwaik
Keyword(s):  
Mass Spectrometry ◽  
Host Cell ◽  
Ubiquitin Ligase ◽  
Legionella Pneumophila ◽  
E3 Ubiquitin Ligase ◽  
Content Type ◽  
Lysine Residues ◽  
Cytosolic Side ◽  
Bona Fide ◽  
F Box Protein

The fate of the polyubiquitinated protein is determined by the lysine linkages involved in the polymerization of the ubiquitin monomers, which has seven lysine residues (K6, K11, K27, K29, K33, K48, and K63). The translocated AnkB effector of the intravacuolar pathogenLegionella pneumophilais a bona fide F-box protein, which is localized to the cytosolic side of theLegionella-containing vacuole (LCV) and is essential for intravacuolar proliferation within macrophages and amoebae. The F-box domain of AnkB interacts with the host SCF1 E3 ubiquitin ligase that triggers the decoration of the LCV with K48-linked polyubiquitinated proteins that are targeted for proteasomal degradation. Here we report that AnkB becomes rapidly polyubiquitinated within the host cell, and this modification is independent of the F-box domain of AnkB, indicating host-mediated polyubiquitination. We show that the AnkB effector interacts specifically with the host E3 ubiquitin ligase Trim21. Mass spectrometry analyses have shown that AnkB is modified by K11-linked polyubiquitination, which has no effect on its stability. This work shows the first example of K11-linked polyubiquitination of a bacterial effector and its interaction with the host Trim21 ubiquitin ligase.

scholarly journals The novel E3 ubiquitin ligase Tiul1 associates with TGIF to target Smad2 for degradation

2004 ◽  
Vol 23 (19) ◽  
pp. 3780-3792 ◽  
Author(s):  
Su Ryeon Seo ◽  
François Lallemand ◽  
Nathalie Ferrand ◽  
Marcia Pessah ◽  
Sébastien L'Hoste ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
E3 Ubiquitin Ligase ◽  
The Novel

scholarly journals Antagonistic Regulation of Circadian Output and Synaptic Development by the E3 Ubiquitin Ligase JETLAG and the DYSCHRONIC-SLOWPOKE Complex

2019 ◽  
Author(s):  
Angelique Lamaze ◽  
James E.C Jepson ◽  
Oghenerukevwe Akpoghiran ◽  
Kyunghee Koh
Keyword(s):  
Neuromuscular Junction ◽  
Ubiquitin Ligase ◽  
Light Exposure ◽  
E3 Ubiquitin Ligase ◽  
List Type ◽  
Cellular Functions ◽  
Synaptic Development ◽  
Channel Expression ◽  
Pdz Protein ◽  
Larval Neuromuscular Junction

SummaryCircadian output genes act downstream of the clock to promote rhythmic changes in behavior and physiology, yet their molecular and cellular functions are not well understood. Here we characterize an interaction between regulators of circadian entrainment, output and synaptic development in Drosophila that influences clock-driven anticipatory increases in morning and evening activity. We previously showed the JETLAG (JET) E3 Ubiquitin ligase resets the clock upon light exposure, while the PDZ protein DYSCHRONIC (DYSC) regulates circadian locomotor output and synaptic development. Surprisingly, we find that JET and DYSC antagonistically regulate synaptic development at the larval neuromuscular junction, and reduced JET activity rescues arrhythmicity of dysc mutants. Consistent with our prior finding that DYSC regulates SLOWPOKE (SLO) potassium channel expression, jet mutations also rescue circadian and synaptic phenotypes in slo mutants. Collectively, our data suggest that JET, DYSC and SLO promote circadian output in part by regulating synaptic morphology.HighlightsLoss of DYSC differentially impacts morning and evening oscillatorsReduced JET activity rescues the dysc and slo arrhythmic phenotypeReduced JET activity causes synaptic defects at the larval NMJJET opposes DYSC and SLO function at the NMJ synapse

scholarly journals The E3 Ubiquitin Ligase Siah-1 Suppresses Avian Reovirus Infection by Targeting p10 for Degradation

2018 ◽  
Vol 92 (6) ◽  
Author(s):  
Xiang Chen ◽  
Zhiyuan He ◽  
Mengjiao Fu ◽  
Yongqiang Wang ◽  
Haiyang Wu ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Ubiquitin Ligase ◽  
Respiratory Diseases ◽  
E3 Ubiquitin Ligase ◽  
Syncytium Formation ◽  
Host Cells ◽  
Avian Reovirus ◽  
Chronic Respiratory Diseases ◽  
Multicomponent Complex ◽  
Seven In Absentia

ABSTRACTAvian reovirus (ARV) causes viral arthritis, chronic respiratory diseases, retarded growth, and malabsorption syndrome. The ARV p10 protein, a viroporin responsible for the induction of cell syncytium formation and apoptosis, is rapidly degraded in host cells. Our previous report demonstrated that cellular lysosome-associated membrane protein 1 (LAMP-1) interacted with p10 and was involved in its degradation. However, the molecular mechanism underlying LAMP-1-mediated p10 degradation remains elusive. We report here that the E3 ubiquitin ligase seven in absentia homolog 1 (Siah-1) is critical for p10 ubiquitylation. Our data show that Siah-1 ubiquitylated p10 and targeted it for proteasome degradation. Furthermore, the ubiquitylation of p10 by Siah-1 required the participation of LAMP-1 by forming a multicomponent complex. Thus, LAMP-1 promotes the proteasomal degradation of p10 via interacting with both p10 and the E3 ligase Siah-1. These data establish a novel host defense mechanism where LAMP-1 serves as a scaffold for both Siah-1 and p10 that allows the E3 ligase targeting p10 for ubiquitylation and degradation to suppress ARV infection.IMPORTANCEAvian reovirus (ARV) is an important poultry pathogen causing viral arthritis, chronic respiratory diseases, retarded growth, and malabsorption syndrome, leading to considerable economic losses to the poultry industry across the globe. The ARV p10 protein is a virulence factor responsible for the induction of cell syncytium formation and apoptosis and is rapidly degraded in host cells. We previously found that cellular lysosome-associated membrane protein 1 (LAMP-1) interacts with p10 and is involved in its degradation. Here we report that the E3 ubiquitin ligase seven in absentia homolog 1 (Siah-1) ubiquitylated p10 and targeted it for proteasomal degradation. Furthermore, the ubiquitylation of p10 by Siah-1 required the participation of LAMP-1 by forming a multicomponent complex. Thus, LAMP-1 serves as an adaptor to allow Siah-1 to target p10 for degradation, thereby suppressing ARV growth in host cells.

scholarly journals PINK1 phosphorylates ubiquitin to activate Parkin E3 ubiquitin ligase activity

2014 ◽  
Vol 205 (2) ◽  
pp. 143-153 ◽  
Author(s):  
Lesley A. Kane ◽  
Michael Lazarou ◽  
Adam I. Fogel ◽  
Yan Li ◽  
Koji Yamano ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Ubiquitin Ligase ◽  
E3 Ubiquitin Ligase ◽  
E3 Ligase ◽  
Feed Forward ◽  
Selective Autophagy ◽  
Ubiquitin Ligase Activity ◽  
In Cells

PINK1 kinase activates the E3 ubiquitin ligase Parkin to induce selective autophagy of damaged mitochondria. However, it has been unclear how PINK1 activates and recruits Parkin to mitochondria. Although PINK1 phosphorylates Parkin, other PINK1 substrates appear to activate Parkin, as the mutation of all serine and threonine residues conserved between Drosophila and human, including Parkin S65, did not wholly impair Parkin translocation to mitochondria. Using mass spectrometry, we discovered that endogenous PINK1 phosphorylated ubiquitin at serine 65, homologous to the site phosphorylated by PINK1 in Parkin’s ubiquitin-like domain. Recombinant TcPINK1 directly phosphorylated ubiquitin and phospho-ubiquitin activated Parkin E3 ubiquitin ligase activity in cell-free assays. In cells, the phosphomimetic ubiquitin mutant S65D bound and activated Parkin. Furthermore, expression of ubiquitin S65A, a mutant that cannot be phosphorylated by PINK1, inhibited Parkin translocation to damaged mitochondria. These results explain a feed-forward mechanism of PINK1-mediated initiation of Parkin E3 ligase activity.

scholarly journals Persistent Mycoplasma genitalium Infection of Human Endocervical Epithelial Cells Elicits Chronic Inflammatory Cytokine Secretion

2012 ◽  
Vol 80 (11) ◽  
pp. 3842-3849 ◽  
Author(s):  
Chris L. McGowin ◽  
Rochelle S. Annan ◽  
Alison J. Quayle ◽  
Sheila J. Greene ◽  
Liang Ma ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Inflammatory Cytokine ◽  
Mycoplasma Genitalium ◽  
Clinical Findings ◽  
Cytokine Secretion ◽  
Host Cells ◽  
Content Type ◽  
Sexually Transmitted ◽  
Host Immune Responses ◽  
Tract Infections

ABSTRACTInfection withMycoplasma genitaliumhas been associated with male and female urogenital disease syndromes, including urethritis, cervicitis, pelvic inflammatory disease (PID), and tubal factor infertility. Basic investigations of mucosal cytotoxicity, microbial persistence, and host immune responses are imperative to understanding these inflammatory urogenital syndromes, particularly in females, considering the potential severity of upper tract infections. Here, we report thatM. genitaliumcan establish long-term infection of human endocervical epithelial cells that results in chronic inflammatory cytokine secretion and increased responsiveness to secondary Toll-like receptor (TLR) stimulation. Using a novel quantitative PCR assay,M. genitaliumwas shown to replicate from 0 to 80 days postinoculation (p.i.), during which at most time points the median ratio ofM. genitaliumorganisms to host cells was ≤10, indicating that low organism burdens are capable of eliciting chronic inflammation in endocervical epithelial cells. This observation is consistent with clinical findings in women. Persistently secreted cytokines predominately consisted of potent chemotactic and/or activating factors for phagocytes, including interleukin-8 (IL-8), monocyte chemotactic protein 1 (MCP-1), and macrophage inflammatory protein 1β (MIP-1β). Despite persistent cytokine elaboration, no host cell cytotoxicity was observed except with superphysiologic loads ofM. genitalium, suggesting that persistent infection occurs with minimal direct damage to the epithelium. However, it is hypothesized that chronic chemokine secretion with leukocyte trafficking to the epithelium could lead to significant inflammatory sequelae. Therefore, persistentM. genitaliuminfection could have important consequences for acquisition and/or pathogenesis of other sexually transmitted infections (STIs) and perhaps explain the positive associations between this organism and human immunodeficiency virus (HIV) shedding.

scholarly journals The E3 Ubiquitin Ligase TBK1 Mediates the Degradation of Multiple Picornavirus VP3 Proteins by Phosphorylation and Ubiquitination

2019 ◽  
Vol 93 (23) ◽  
Author(s):  
Dan Li ◽  
Wenping Yang ◽  
Jingjing Ren ◽  
Yi Ru ◽  
Keshan Zhang ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Molecular Mechanisms ◽  
E3 Ubiquitin Ligase ◽  
Adaptor Protein ◽  
Ubiquitin Ligase Activity ◽  
Lysine Residues ◽  
Innate Antiviral Immunity

ABSTRACT TANK-binding kinase 1 (TBK1) is essential for interferon beta (IFN-β) production and innate antiviral immunity. However, other, additional functions of TBK1 have remained elusive. Here, we showed that TBK1 is an E3 ubiquitin ligase that undergoes self-ubiquitylation in vitro in the presence of the E2 enzyme UbcH5c. Further evidence showed that TBK1 could also be self-ubiquitylated in vivo. Importantly, multiple picornavirus VP3 proteins were degraded by TBK1 through its kinase and E3 ubiquitin ligase activity. Mechanistically, TBK1 phosphorylated multiple picornavirus VP3 proteins at serine residues and ubiquitinated them via K63-linked ubiquitination at lysine residues. In addition, the C426 and C605 residues of TBK1 were not essential for TBK1 innate immunity activity; however, these residues were required for degradation of multiple picornavirus VP3 proteins and for its E3 ubiquitin ligase activity. Hence, our findings identified a novel role of TBK1 in regulating the virus life cycle and provided new insights into the molecular mechanisms of TBK1-mediated antiviral response. IMPORTANCE TBK1 is an important adaptor protein required for innate immune response to viruses, but its other functions were unknown. In this study, we found that TBK1 is an E3 ubiquitin ligase that undergoes self-ubiquitylation in vitro in the presence of the E2 enzyme UbcH5c. In addition, multiple picornavirus VP3 proteins were degraded by TBK1 through its kinase and E3 ubiquitin ligase activity. Our report provides evidence that TBK1 plays a role in viral protein degradation.

scholarly journals The novel multi-cytokine inhibitor TO-207 specifically inhibits pro-inflammatory cytokine secretion in monocytes without affecting the killing ability of CAR T cells

PLoS ONE ◽  
2020 ◽  
Vol 15 (4) ◽  
pp. e0231896 ◽  
Author(s):  
Muneyoshi Futami ◽  
Keisuke Suzuki ◽  
Satomi Kato ◽  
Saori Ohmae ◽  
Yoshio Tahara ◽  
...  
Keyword(s):  
T Cells ◽  
Inflammatory Cytokine ◽  
Cytokine Secretion ◽  
The Novel ◽  
Car T Cells ◽  
Car T ◽  
Cytokine Inhibitor

scholarly journals Ubiquitin systems mark pathogen-containing vacuoles as targets for host defense by guanylate binding proteins

2015 ◽  
Vol 112 (41) ◽  
pp. E5628-E5637 ◽  
Author(s):  
Arun K. Haldar ◽  
Clémence Foltz ◽  
Ryan Finethy ◽  
Anthony S. Piro ◽  
Eric M. Feeley ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Microbial Growth ◽  
E3 Ubiquitin Ligase ◽  
Host Cells ◽  
Tnf Receptor ◽  
Host Proteins ◽  
Growth And Survival ◽  
Guanylate Binding Protein ◽  
Rhoptry Protein ◽  
Necrosis Factor

Many microbes create and maintain pathogen-containing vacuoles (PVs) as an intracellular niche permissive for microbial growth and survival. The destruction of PVs by IFNγ-inducible guanylate binding protein (GBP) and immunity-related GTPase (IRG) host proteins is central to a successful immune response directed against numerous PV-resident pathogens. However, the mechanism by which IRGs and GBPs cooperatively detect and destroy PVs is unclear. We find that host cell priming with IFNγ prompts IRG-dependent association of Toxoplasma- and Chlamydia-containing vacuoles with ubiquitin through regulated translocation of the E3 ubiquitin ligase tumor necrosis factor (TNF) receptor associated factor 6 (TRAF6). This initial ubiquitin labeling elicits p62-mediated escort and deposition of GBPs to PVs, thereby conferring cell-autonomous immunity. Hypervirulent strains of Toxoplasma gondii evade this process via specific rhoptry protein kinases that inhibit IRG function, resulting in blockage of downstream PV ubiquitination and GBP delivery. Our results define a ubiquitin-centered mechanism by which host cells deliver GBPs to PVs and explain how hypervirulent parasites evade GBP-mediated immunity.

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