scholarly journals IKKγ/NEMO Is Required to Confer Antimicrobial Innate Immune Responses in the Yellow Mealworm, Tenebrio Molitor

2020 ◽  
Vol 21 (18) ◽  
pp. 6734
Author(s):  
Hye Jin Ko ◽  
Yong Hun Jo ◽  
Bharat Bhusan Patnaik ◽  
Ki Beom Park ◽  
Chang Eun Kim ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Leucine Zipper ◽  
Fat Body ◽  
Coiled Coil ◽  
Tenebrio Molitor ◽  
Innate Immune ◽  
Regulatory Subunit ◽  
Iκb Kinase ◽  
Body Tissues ◽  
Ikk Complex

IKKγ/NEMO is the regulatory subunit of the IκB kinase (IKK) complex, which regulates the NF-κB signaling pathway. Within the IKK complex, IKKγ/NEMO is the non-catalytic subunit, whereas IKKα and IKKβ are the structurally related catalytic subunits. In this study, TmIKKγ was screened from the Tenebrio molitor RNA-Seq database and functionally characterized using RNAi screening for its role in regulating T. molitor antimicrobial peptide (AMP) genes after microbial challenges. The TmIKKγ transcript is 1521 bp that putatively encodes a polypeptide of 506 amino acid residues. TmIKKγ contains a NF-κB essential modulator (NEMO) and a leucine zipper domain of coiled coil region 2 (LZCC2). A phylogenetic analysis confirmed its homology to the red flour beetle, Tribolium castaneum IKKγ (TcIKKγ). The expression of TmIKKγ mRNA showed that it might function in diverse tissues of the insect, with a higher expression in the hemocytes and the fat body of the late-instar larvae. TmIKKγ mRNA expression was induced by Escherichia coli, Staphylococcus aureus, and Candida albicans challenges in the whole larvae and in tissues such as the hemocytes, gut and fat body. The knockdown of TmIKKγ mRNA significantly reduced the survival of the larvae after microbial challenges. Furthermore, we investigated the tissue-specific induction patterns of fourteen T. molitor AMP genes in TmIKKγ mRNA-silenced individuals after microbial challenges. In general, the mRNA expression of TmTenecin1, -2, and -4; TmDefensin1 and -2; TmColeoptericin1 and 2; and TmAttacin1a, 1b, and 2 were found to be downregulated in the hemocytes, gut, and fat body tissues in the TmIKKγ-silenced individuals after microbial challenges. Under similar conditions, TmRelish (NF-κB transcription factor) mRNA was also found to be downregulated. Thus, TmIKKγ is an important factor in the antimicrobial innate immune response of T. molitor.

scholarly journals NEMO oligomerization and its ubiquitin-binding properties

2009 ◽  
Vol 421 (2) ◽  
pp. 243-251 ◽  
Author(s):  
Frank J. Ivins ◽  
Mark G. Montgomery ◽  
Susan J. M. Smith ◽  
Aylin C. Morris-Davies ◽  
Ian A. Taylor ◽  
...  
Keyword(s):  
Leucine Zipper ◽  
Target Genes ◽  
Nuclear Translocation ◽  
Coiled Coil ◽  
Nuclear Factor Κb ◽  
Regulatory Subunit ◽  
Activation Process ◽  
Oligomeric State ◽  
Coiled Coil Region ◽  
Ikk Complex

The IKK [IκB (inhibitory κB) kinase] complex is a key regulatory component of NF-κB (nuclear factor κB) activation and is responsible for mediating the degradation of IκB, thereby allowing nuclear translocation of NF-κB and transcription of target genes. NEMO (NF-κB essential modulator), the regulatory subunit of the IKK complex, plays a pivotal role in this process by integrating upstream signals, in particular the recognition of polyubiquitin chains, and relaying these to the activation of IKKα and IKKβ, the catalytic subunits of the IKK complex. The oligomeric state of NEMO is controversial and the mechanism by which it regulates activation of the IKK complex is poorly understood. Using a combination of hydrodynamic techniques we now show that apo-NEMO is a highly elongated, dimeric protein that is in weak equilibrium with a tetrameric assembly. Interaction with peptides derived from IKKβ disrupts formation of the tetrameric NEMO complex, indicating that interaction with IKKα and IKKβ and tetramerization are mutually exclusive. Furthermore, we show that NEMO binds to linear di-ubiquitin with a stoichiometry of one molecule of di-ubiquitin per NEMO dimer. This stoichiometry is preserved in a construct comprising the second coiled-coil region and the leucine zipper and in one that essentially spans the full-length protein. However, our data show that at high di-ubiquitin concentrations a second weaker binding site becomes apparent, implying that two different NEMO–di-ubiquitin complexes are formed during the IKK activation process. We propose that the role of these two complexes is to provide a threshold for activation, thereby ensuring sufficient specificity during NF-κB signalling.

scholarly journals Hepatitis B e Antigen Inhibits NF-κB Activity by Interrupting K63-Linked Ubiquitination of NEMO

2018 ◽  
Vol 93 (2) ◽  
Author(s):  
Yuan Wang ◽  
Lei Cui ◽  
Guifang Yang ◽  
Jianbo Zhan ◽  
Liang Guo ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Signaling Pathway ◽  
Molecular Mechanism ◽  
Virus Replication ◽  
Innate Immune ◽  
Regulatory Subunit ◽  
Hepatitis B E Antigen ◽  
Iκb Kinase ◽  
B Virus

ABSTRACTViruses have adopted diverse strategies to suppress antiviral responses. Hepatitis B virus (HBV), a virus that is prevalent worldwide, manipulates the host’s innate immune system to evade scavenging. It is reported that the hepatitis B e antigen (HBeAg) can interfere with NF-κB activity, which then leads to high viral loads, while HBV with the G1896A mutation remains infectious without the production of HBeAg but can induce more severe proinflammatory response and liver damage. The aim of current work was to study the molecular mechanism by which HBeAg suppresses interleukin-1β (IL-1β)-stimulated NF-κB activity, which leads to the suppression of the innate immune responses to HBV infection. Our study revealed that HBeAg could interact with NEMO, a regulatory subunit associated with IκB kinase, which regulates the activation of NF-κB. HBeAg suppressed the IL-1β-induced tumor necrosis factor (TNF)-associated factor 6 (TRAF6)-dependent K63-linked ubiquitination of NEMO, thereby downregulating NF-κB activity and promoting virus replication. We further demonstrated the inhibitory effect of HBeAg on the NF-κB signaling pathway using primary human hepatocytes, HBV-infected HepG2-NTCP cells, and clinical liver samples. Our study reveals a molecular mechanism whereby HBeAg suppresses IL-1β-induced NF-κB activation by decreasing the TRAF6-dependent K63-linked ubiquitination of NEMO, which may thereby enhance HBV replication and promote a persistent infection.IMPORTANCEThe role of HBeAg in inflammatory responses during the infection of hepatitis B virus (HBV) is not fully understood, and several previous reports with regard to the NF-κB pathway are controversial. In this study, we showed that HBeAg could suppress both Toll-like receptor 2 (TLR2)- and IL-1β-induced activation of NF-κB in cells and clinical samples, and we further revealed novel molecular mechanisms. We found that HBeAg can associate with NEMO, the regulatory subunit for IκB kinase (IKK) that controls the NF-κB signaling pathway, and thereby inhibits TRAF6-mediated K63-linked ubiquitination of NEMO, resulting in downregulation of NF-κB activity and promotion of virus replication. In contrast, the HBeAg-negative HBV mutant can induce higher levels of NF-κB activity. These results are important for understanding the HBV-induced pathogenesis of chronic hepatitis and indicate that different clinical measures should be considered to treat HBeAg-positive and HBeAg-negative infections. Our findings represent a conceptual advance in HBV-related suppression of NF-κB signaling.

scholarly journals IκB Kinase Signaling Is Essential for Maintenance of Mature B Cells

2002 ◽  
Vol 196 (6) ◽  
pp. 743-752 ◽  
Author(s):  
Manolis Pasparakis ◽  
Marc Schmidt-Supprian ◽  
Klaus Rajewsky
Keyword(s):  
B Cells ◽  
Regulatory Subunit ◽  
Cellular Survival ◽  
Adult Mice ◽  
Iκb Kinase ◽  
Ikk Complex ◽  
Conditional Gene Targeting ◽  
B Lineage ◽  
Deficient Mice

Nuclear factor (NF)-κB proteins play crucial roles in immune responses and cellular survival. Activation of NF-κB is mediated by the IκB kinase (IKK) complex, which is composed of two kinases, IKK1 and IKK2, and a regulatory subunit termed NF-κB essential modulator (NEMO). IKK2- and NEMO-deficient mice die at early embryonic stages. We therefore used conditional gene targeting to evaluate the role of these proteins in B cells in adult mice. B lineage–specific disruption of either IKK signaling by deletion of NEMO, or of IKK2-specific signals by ablation of IKK2 activity leads to the disappearance of mature B lymphocytes. We conclude that maintenance of mature B cells depends on IKK-mediated activation of NF-κB.

scholarly journals The IKKβ Subunit of IκB Kinase (IKK) is Essential for Nuclear Factor κB Activation and Prevention of Apoptosis

1999 ◽  
Vol 189 (11) ◽  
pp. 1839-1845 ◽  
Author(s):  
Zhi-Wei Li ◽  
Wenming Chu ◽  
Yinling Hu ◽  
Mireille Delhase ◽  
Tom Deerinck ◽  
...  
Keyword(s):  
Interleukin 1 ◽  
Nuclear Factor Κb ◽  
Regulatory Subunit ◽  
Nuclear Factor ◽  
Catalytic Subunits ◽  
Iκb Kinase ◽  
Ikk Complex ◽  
Factor Α ◽  
Necrosis Factor

The IκB kinase (IKK) complex is composed of three subunits, IKKα, IKKβ, and IKKγ (NEMO). While IKKα and IKKβ are highly similar catalytic subunits, both capable of IκB phosphorylation in vitro, IKKγ is a regulatory subunit. Previous biochemical and genetic analyses have indicated that despite their similar structures and in vitro kinase activities, IKKα and IKKβ have distinct functions. Surprisingly, disruption of the Ikkα locus did not abolish activation of IKK by proinflammatory stimuli and resulted in only a small decrease in nuclear factor (NF)-κB activation. Now we describe the pathophysiological consequence of disruption of the Ikkβ locus. IKKβ-deficient mice die at mid-gestation from uncontrolled liver apoptosis, a phenotype that is remarkably similar to that of mice deficient in both the RelA (p65) and NF-κB1 (p50/p105) subunits of NF-κB. Accordingly, IKKβ-deficient cells are defective in activation of IKK and NF-κB in response to either tumor necrosis factor α or interleukin 1. Thus IKKβ, but not IKKα, plays the major role in IKK activation and induction of NF-κB activity. In the absence of IKKβ, IKKα is unresponsive to IKK activators.

scholarly journals Roles of GSK-3 and β-Catenin in Antiviral Innate Immune Sensing of Nucleic Acids

Cells ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 897 ◽  
Author(s):  
Alexandre Marineau ◽  
Kashif Aziz Khan ◽  
Marc J. Servant
Keyword(s):  
Immune Response ◽  
Innate Immune Response ◽  
Cellular Response ◽  
Glycogen Synthase ◽  
Innate Immune ◽  
Type I ◽  
Resting Cells ◽  
Iκb Kinase ◽  
Ikk Complex ◽  
Rna And Dna

The rapid activation of the type I interferon (IFN) antiviral innate immune response relies on ubiquitously expressed RNA and DNA sensors. Once engaged, these nucleotide-sensing receptors use distinct signaling modules for the rapid and robust activation of mitogen-activated protein kinases (MAPKs), the IκB kinase (IKK) complex, and the IKK-related kinases IKKε and TANK-binding kinase 1 (TBK1), leading to the subsequent activation of the activator protein 1 (AP1), nuclear factor-kappa B (NF-κB), and IFN regulatory factor 3 (IRF3) transcription factors, respectively. They, in turn, induce immunomodulatory genes, allowing for a rapid antiviral cellular response. Unlike the MAPKs, the IKK complex and the IKK-related kinases, ubiquitously expressed glycogen synthase kinase 3 (GSK-3) α and β isoforms are active in unstimulated resting cells and are involved in the constitutive turnover of β-catenin, a transcriptional coactivator involved in cell proliferation, differentiation, and lineage commitment. Interestingly, studies have demonstrated the regulatory roles of both GSK-3 and β-catenin in type I IFN antiviral innate immune response, particularly affecting the activation of IRF3. In this review, we summarize current knowledge on the mechanisms by which GSK-3 and β-catenin control the antiviral innate immune response to RNA and DNA virus infections.

scholarly journals Tetrameric Oligomerization of IκB Kinase γ (IKKγ) Is Obligatory for IKK Complex Activity and NF-κB Activation

2003 ◽  
Vol 23 (6) ◽  
pp. 2029-2041 ◽  
Author(s):  
Sebastian Tegethoff ◽  
Joachim Behlke ◽  
Claus Scheidereit
Keyword(s):  
Kinase Activity ◽  
Coiled Coil ◽  
Complex Activity ◽  
Catalytic Subunits ◽  
Iκb Kinase ◽  
293 Cells ◽  
Activation Of Transcription ◽  
Ikk Complex ◽  
Spatial Positioning ◽  
Oligomerization Domain

ABSTRACT The IκB kinase (IKK) complex mediates activation of transcription factor NF-κB by phosphorylation of IκB proteins. Its catalytic subunits, IKKα and IKKβ, require association with the regulatory IKKγ (NEMO) component to gain full basal and inducible kinase activity. However, the oligomeric composition of the IKK complex and its regulation by IKKγ are poorly understood. We show here that IKKγ predominantly forms tetramers and interacts with IKKα or IKKβ in this state. We propose that tetramerization is accomplished by a prerequisite dimerization through a C-terminal coiled-coil minimal oligomerization domain (MOD). This is followed by dimerization of the dimers with their N-terminal sequences. Tetrameric IKKγ sequesters four kinase molecules, yielding a γ4(α/β)4 stoichiometry. Deletion of the MOD leads to loss of tetramerization and of phosphorylation of IKKβ and IKKγ, although the kinase can still interact with the resultant IKKγ monomers and dimers. Likewise, MOD-mediated IKKγ tetramerization is required to enhance IKKβ kinase activity when overexpressed in 293 cells and to reconstitute a lipopolysaccharide-responsive IKK complex in pre-B cells. These data thus suggest that IKKγ tetramerization enforces a spatial positioning of two kinase dimers to facilitate transautophosphorylation and activation.

scholarly journals FAF1 Suppresses IκB Kinase (IKK) Activation by Disrupting the IKK Complex Assembly

2007 ◽  
Vol 282 (38) ◽  
pp. 27572-27577 ◽  
Author(s):  
Min-Young Park ◽  
Ji-hyun Moon ◽  
Ki-Sung Lee ◽  
Hye-In Choi ◽  
Jongkyeong Chung ◽  
...  
Keyword(s):  
Leucine Zipper ◽  
Physical Interaction ◽  
Inhibitory Mechanism ◽  
Complex Assembly ◽  
Downstream Signaling ◽  
Leucine Zipper Domain ◽  
Iκb Kinase ◽  
Ikk Complex ◽  
Factor Α ◽  
Necrosis Factor

This study presents a molecular inhibitory mechanism by Fas-associated factor 1 (FAF1) on IκB kinase (IKK) activation, where divergent NF-κB-activating stimuli converge. FAF1 interacts with IKKβ in response to proinflammatory stimuli (such as tumor necrosis factor-α, interleukin-1β, and lipopolysaccharide) and suppresses IKK activation. Interaction of the leucine-zipper domain of IKKβ with FAF1 affected the IKK heterocomplex (IKKα/β) and homocomplex (IKKα/α, IKKβ/β) formations and attenuated IKKγ recruitment to IKKβ. Overexpression of FAF1 reduced the level of IKKβ activity, whereas FAF1 depletion increased the activity. These results indicate that FAF1 inhibits IKK activation and its downstream signaling by interrupting the IKK complex assembly through physical interaction with IKKβ. Taken together, FAF1 robustly suppresses NF-κB activation through the inhibition of IKK activation in combination with previously reported cytoplasmic retention of NF-κB p65 (Park, M. Y., Jang, H. D., Lee, S. Y., Lee, K. J., and Kim, E. (2004) J. Biol. Chem. 279, 2544–2549). Such redundant suppression would prevent inadvertent activation of the NF-κB pathway.

Mitochondrial reactive oxygen species enable proinflammatory signaling through disulfide linkage of NEMO

2019 ◽  
Vol 12 (568) ◽  
pp. eaar5926 ◽  
Author(s):  
Marc Herb ◽  
Alexander Gluschko ◽  
Katja Wiegmann ◽  
Alina Farid ◽  
Anne Wolf ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Reactive Oxygen ◽  
Regulatory Subunit ◽  
Oxygen Species ◽  
Proinflammatory Response ◽  
Disulfide Linkage ◽  
Macrophage Response ◽  
Major Function ◽  
Iκb Kinase ◽  
Ikk Complex

A major function of macrophages during infection is initiation of the proinflammatory response, leading to the secretion of cytokines that help to orchestrate the immune response. Here, we identify reactive oxygen species (ROS) as crucial mediators of proinflammatory signaling leading to cytokine secretion in Listeria monocytogenes–infected macrophages. ROS produced by NADPH oxidases (Noxes), such as Nox2, are key components of the macrophage response to invading pathogens; however, our data show that the ROS that mediated proinflammatory signaling were produced by mitochondria (mtROS). We identified the inhibitor of κB (IκB) kinase (IKK) complex regulatory subunit NEMO [nuclear factor κB (NF-κB) essential modulator] as a target for mtROS. Specifically, mtROS induced intermolecular covalent linkage of NEMO through disulfide bonds formed by Cys54 and Cys347, which was essential for activation of the IKK complex and subsequent signaling through the extracellular signal–regulated protein kinases 1 and 2 (ERK1/2) and NF-κB pathways that eventually led to the secretion of proinflammatory cytokines. We thus identify mtROS-dependent disulfide linkage of NEMO as an essential regulatory step of the proinflammatory response of macrophages to bacterial infection.

scholarly journals Annexin-A1 SUMOylation regulates microglial polarization after cerebral ischemia by modulating IKKα stability via selective autophagy

2021 ◽  
Vol 7 (4) ◽  
pp. eabc5539
Author(s):  
Xing Li ◽  
Qian Xia ◽  
Meng Mao ◽  
Huijuan Zhou ◽  
Lu Zheng ◽  
...  
Keyword(s):  
Cerebral Ischemia ◽  
Underlying Mechanism ◽  
Nuclear Factor Κb ◽  
Annexin A1 ◽  
Selective Autophagy ◽  
Microglial Polarization ◽  
Proinflammatory Mediators ◽  
Therapeutic Benefits ◽  
Iκb Kinase ◽  
Ikk Complex

Annexin-A1 (ANXA1) has recently been proposed to play a role in microglial activation after brain ischemia, but the underlying mechanism remains poorly understood. Here, we demonstrated that ANXA1 is modified by SUMOylation, and SUMOylated ANXA1 could promote the beneficial phenotype polarization of microglia. Mechanistically, SUMOylated ANXA1 suppressed nuclear factor κB activation and the production of proinflammatory mediators. Further study revealed that SUMOylated ANXA1 targeted the IκB kinase (IKK) complex and selectively enhanced IKKα degradation. Simultaneously, we detected that SUMOylated ANXA1 facilitated the interaction between IKKα and NBR1 to promote IKKα degradation through selective autophagy. Further work revealed that the overexpression of SUMOylated ANXA1 in microglia/macrophages resulted in marked improvement in neurological function in a mouse model of cerebral ischemia. Collectively, our study demonstrates a previously unidentified mechanism whereby SUMOylated ANXA1 regulates microglial polarization and strongly indicates that up-regulation of ANXA1 SUMOylation in microglia may provide therapeutic benefits for cerebral ischemia.

scholarly journals The α and β Subunits of IκB Kinase (IKK) Mediate TRAF2-Dependent IKK Recruitment to Tumor Necrosis Factor (TNF) Receptor 1 in Response to TNF

2001 ◽  
Vol 21 (12) ◽  
pp. 3986-3994 ◽  
Author(s):  
Anne Devin ◽  
Yong Lin ◽  
Shoji Yamaoka ◽  
Zhiwei Li ◽  
Michael Karin ◽  
...  
Keyword(s):  
Tumor Necrosis Factor ◽  
Tumor Necrosis ◽  
Leucine Zipper ◽  
Nuclear Translocation ◽  
Ring Finger ◽  
Tnf Receptor ◽  
Death Domain ◽  
Iκb Kinase ◽  
Cytokine Tumor Necrosis Factor ◽  
Necrosis Factor

ABSTRACT The activation of IκB kinase (IKK) is a key step in the nuclear translocation of the transcription factor NF-κB. IKK is a complex composed of three subunits: IKKα, IKKβ, and IKKγ (also called NEMO). In response to the proinflammatory cytokine tumor necrosis factor (TNF), IKK is activated after being recruited to the TNF receptor 1 (TNF-R1) complex via TNF receptor-associated factor 2 (TRAF2). We found that the IKKα and IKKβ catalytic subunits are required for IKK-TRAF2 interaction. This interaction occurs through the leucine zipper motif common to IKKα, IKKβ, and the RING finger domain of TRAF2, and either IKKα or IKKβ alone is sufficient for the recruitment of IKK to TNF-R1. Importantly, IKKγ is not essential for TNF-induced IKK recruitment to TNF-R1, as this occurs efficiently in IKKγ-deficient cells. Using TRAF2−/− cells, we demonstrated that the TNF-induced interaction between IKKγ and the death domain kinase RIP is TRAF2 dependent and that one possible function of this interaction is to stabilize the IKK complex when it interacts with TRAF2.

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