Regulation of TRIF-mediated innate immune response by K27-linked polyubiquitination and deubiquitination

Abstract TIR domain-containing adaptor inducing interferon-β (TRIF) is an essential adaptor protein required for innate immune responses mediated by Toll-like receptor (TLR) 3- and TLR4. Here we identify USP19 as a negative regulator of TLR3/4-mediated signaling. USP19 deficiency increases the production of type I interferons (IFN) and proinflammatory cytokines induced by poly(I:C) or LPS in vitro and in vivo. Usp19-/- mice have more serious inflammation after poly(I:C) or LPS treatment, and are more susceptible to inflammatory damages and death following Salmonella typhimurium infection. Mechanistically, USP19 interacts with TRIF and catalyzes the removal of TRIF K27-linked polyubiquitin moieties, thereby impairing the recruitment of TRIF to TLR3/4. In addition, the RING E3 ubiquitin ligase complex Cullin-3-Rbx1-KCTD10 catalyzes K27-linked polyubiquitination of TRIF at K523, and deficiency of this complex inhibits TLR3/4-mediated innate immune signaling. Our findings thus reveal TRIF K27-linked polyubiquitination and deubiquitination as a critical regulatory mechanism of TLR3/4-mediated innate immune responses.

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PPP6C Negatively Regulates STING-Dependent Innate Immune Responses

mBio ◽

10.1128/mbio.01728-20 ◽

2020 ◽

Vol 11 (4) ◽

Author(s):

Guoxin Ni ◽

Zhe Ma ◽

Jason P. Wong ◽

Zhigang Zhang ◽

Emily Cousins ◽

...

Keyword(s):

Immune Responses ◽

Adaptor Protein ◽

Innate Immune ◽

Negative Regulator ◽

Host Cells ◽

Type I ◽

Innate Immune Responses ◽

Pathogen Invasion ◽

Major Player ◽

Sting Pathway

ABSTRACT Stimulator of interferon genes (STING) is an essential adaptor protein of the innate DNA-sensing signaling pathway, which recognizes genomic DNA from invading pathogens to establish antiviral responses in host cells. STING activity is tightly regulated by several posttranslational modifications, including phosphorylation. However, specifically how the phosphorylation status of STING is modulated by kinases and phosphatases remains to be fully elucidated. In this study, we identified protein phosphatase 6 catalytic subunit (PPP6C) as a binding partner of Kaposi’s sarcoma-associated herpesvirus (KSHV) open reading frame 48 (ORF48), which is a negative regulator of the cyclic GMP-AMP synthase (cGAS)-STING pathway. PPP6C depletion enhances double-stranded DNA (dsDNA)-induced and 5′ppp double-stranded RNA (dsRNA)-induced but not poly(I:C)-induced innate immune responses. PPP6C negatively regulates dsDNA-induced IRF3 activation but not NF-κB activation. Deficiency of PPP6C greatly inhibits the replication of herpes simplex virus 1 (HSV-1) and vesicular stomatitis virus (VSV) as well as the reactivation of KSHV, due to increased type I interferon production. We further demonstrated that PPP6C interacts with STING and that loss of PPP6C enhances STING phosphorylation. These data demonstrate the important role of PPP6C in regulating STING phosphorylation and activation, which provides an additional mechanism by which the host responds to viral infection. IMPORTANCE Cytosolic DNA, which usually comes from invading microbes, is a dangerous signal to the host. The cGAS-STING pathway is the major player that detects cytosolic DNA and then evokes the innate immune response. As an adaptor protein, STING plays a central role in controlling activation of the cGAS-STING pathway. Although transient activation of STING is essential to trigger the host defense during pathogen invasion, chronic STING activation has been shown to be associated with several autoinflammatory diseases. Here, we report that PPP6C negatively regulates the cGAS-STING pathway by removing STING phosphorylation, which is required for its activation. Dephosphorylation of STING by PPP6C helps prevent the sustained production of STING-dependent cytokines, which would otherwise lead to severe autoimmune disorders. This work provides additional mechanisms on the regulation of STING activity and might facilitate the development of novel therapeutics designed to prevent a variety of autoinflammatory disorders.

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An In Vitro Assessment of Immunostimulatory Responses to Ten Model Innate Immune Response Modulating Impurities (IIRMIs) and Peptide Drug Product, Teriparatide

Molecules ◽

10.3390/molecules26247461 ◽

2021 ◽

Vol 26 (24) ◽

pp. 7461

Author(s):

Claire K. Holley ◽

Edward Cedrone ◽

Duncan Donohue ◽

Barry W. Neun ◽

Daniela Verthelyi ◽

...

Keyword(s):

Immune Response ◽

Immune Responses ◽

Innate Immune Response ◽

Innate Immune ◽

Cytokine Secretion ◽

Poly I:C ◽

In Vitro Assays ◽

Innate Immune Responses ◽

Vitro Assay

Understanding, predicting, and minimizing the immunogenicity of peptide-based therapeutics are of paramount importance for ensuring the safety and efficacy of these products. The so-called anti-drug antibodies (ADA) may have various clinical consequences, including but not limited to the alteration in the product’s distribution, biological activity, and clearance profiles. The immunogenicity of biotherapeutics can be influenced by immunostimulation triggered by the presence of innate immune response modulating impurities (IIRMIs) inadvertently introduced during the manufacturing process. Herein, we evaluate the applicability of several in vitro assays (i.e., complement activation, leukocyte proliferation, and cytokine secretion) for the screening of innate immune responses induced by ten common IIRMIs (Bacillus subtilis flagellin, FSL-1, zymosan, ODN2006, poly(I:C) HMW, poly(I:C) LMW, CLO75, MDP, ODN2216, and Escherichia coli O111:B4 LPS), and a model biotherapeutic Forteo™ (teriparatide). Our study identifies cytokine secretion from healthy human donor peripheral blood mononuclear cells (PBMC) as a sensitive method for the in vitro monitoring of innate immune responses to individual IIRMIs and teriparatide (TP). We identify signature cytokines, evaluate both broad and narrow multiplex cytokine panels, and discuss how the assay logistics influence the performance of this in vitro assay.

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Enhancing innate antiviral immune responses in rainbow trout by double stranded RNA delivered with cationic phytoglycogen nanoparticles

Scientific Reports ◽

10.1038/s41598-019-49931-2 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 4

Author(s):

Tamiru N. Alkie ◽

Jondavid de Jong ◽

Kristof Jenik ◽

Karl M. Klinger ◽

Stephanie J. DeWitte-Orr

Keyword(s):

Rainbow Trout ◽

Immune Responses ◽

Cell Line ◽

Culture Conditions ◽

Type I Interferons ◽

Poly I:C ◽

Synthetic Analogue ◽

Type I ◽

Viral Dsrna

Abstract Innate immunity is induced when pathogen-associated molecular patterns (PAMPs) bind host pattern recognition receptors (PRRs). Polyinosinic:polycytidylic acid [poly(I:C)] is a synthetic analogue of viral dsRNA that acts as a PAMP, inducing type I interferons (IFNs) in vertebrates. In the present study, the immunostimulatory effects of high molecular weight (HMW) poly(I:C) in rainbow trout cells were measured when bound to a cationic phytoglycogen nanoparticle (Nano-HMW). The physical characteristics of the nanoparticle itself, when bound to different lengths of dsRNA and when cell associated was evaluated. Optimal concentration and timing for innate immune stimulation was measured using the RTG-P1 reporter cell line. The immunostimulatory effects of HMW poly (I:C) was compared to Nano-HMW in vitro using the RTgutGC cell line cultured in a conventional monolayer or a transwell culture system. The ability of an activated intestinal epithelium to transmit an antiviral signal to macrophages was evaluated using a co-culture of RTgutGC cells and RTSll (a monocyte/macrophage cell). In all culture conditions, Nano-HMW was a more effective inducer of IFN-related antiviral immune responses compared to HMW poly (I:C) alone. This study introduces the use of cationic phytoglycogen nanoparticles as a novel delivery system for immunomodulatory molecules to enhance immune responses in aquatic vertebrates.

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High-Resolution Profiling of Innate Immune Responses by Porcine Dendritic Cell Subsets in vitro and in vivo

Frontiers in Immunology ◽

10.3389/fimmu.2020.01429 ◽

2020 ◽

Vol 11 ◽

Cited By ~ 1

Author(s):

Gaël Auray ◽

Stephanie C. Talker ◽

Irene Keller ◽

Sylvie Python ◽

Markus Gerber ◽

...

Keyword(s):

Dendritic Cell ◽

High Resolution ◽

Immune Responses ◽

Innate Immune ◽

Innate Immune Responses ◽

Dendritic Cell Subsets

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194 A Non-Gluten Component of Wheat is a Strong Stimulator of Innate Immune Responses In Vitro and In Vivo and Acts via Toll Like Receptor 4

Gastroenterology ◽

10.1016/s0016-5085(10)60165-5 ◽

2010 ◽

Vol 138 (5) ◽

pp. S-36

Author(s):

Yvonne Junker ◽

Donatella Barisani ◽

Daniel A. Leffler ◽

Towia Libermann ◽

Simon T. Dillon ◽

...

Keyword(s):

Immune Responses ◽

Innate Immune ◽

Innate Immune Responses ◽

Toll Like Receptor ◽

Toll Like Receptor 4

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Cytosolic DNA sensor-initiated innate immune responses in mouse ovarian granulosa cells

Reproduction ◽

10.1530/rep-16-0674 ◽

2017 ◽

Vol 153 (6) ◽

pp. 821-834 ◽

Cited By ~ 4

Author(s):

Keqin Yan ◽

Dingqing Feng ◽

Jing Liang ◽

Qing Wang ◽

Lin Deng ◽

...

Keyword(s):

Immune Responses ◽

Granulosa Cells ◽

Innate Immune ◽

Type I Interferons ◽

Endocrine Function ◽

Type I ◽

Dna Sensor ◽

Innate Immune Responses ◽

Oligoadenylate Synthetase ◽

Ovarian Granulosa Cells

Viral infections of the ovary may perturb ovarian functions. However, the mechanisms underlying innate immune responses in the ovary are poorly understood. The present study demonstrates that cytosolic viral DNA sensor signaling initiates the innate immune response in mouse ovarian granulosa cells and affects endocrine function. The cytosolic DNA sensors p204 and cGAS and their common signaling adaptor stimulator of interferon (IFN) genes (STING) were constitutively expressed in granulosa cells. Transfection with VACV70, a synthetic vaccinia virus (VACV) DNA analog, induced the expression of type I interferons (IFNA/B) and major inflammatory cytokines (TNFA and IL6) through IRF3 and NF-κB activation respectively. Moreover, several IFN-inducible antiviral proteins, including 2′,5′-oligoadenylate synthetase, IFN-stimulating gene 15 and Mx GTPase 1, were also induced by VACV70 transfection. The innate immune responses in granulosa cells were significantly reduced by the transfection of specific small-interfering RNAs targeting p204, cGas or Sting. Notably, the VACV70-triggered innate immune responses affected steroidogenesis in vivo and in vitro. The data presented in this study describe the mechanism underlying ovarian immune responses to viral infection.

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How the Innate Immune DNA Sensing cGAS–STING Pathway Is Involved in Autophagy

International Journal of Molecular Sciences ◽

10.3390/ijms222413232 ◽

2021 ◽

Vol 22 (24) ◽

pp. 13232

Author(s):

Wanglong Zheng ◽

Nengwen Xia ◽

Jiajia Zhang ◽

Nanhua Chen ◽

François Meurens ◽

...

Keyword(s):

Immune Responses ◽

Inflammatory Responses ◽

Complex Interaction ◽

Innate Immune ◽

Type I Interferons ◽

Type I ◽

Innate Immune Responses ◽

Homeostatic Process ◽

Sting Pathway ◽

Dna Complex

The cGAS–STING pathway is a key component of the innate immune system and exerts crucial roles in the detection of cytosolic DNA and invading pathogens. Accumulating evidence suggests that the intrinsic cGAS–STING pathway not only facilitates the production of type I interferons (IFN-I) and inflammatory responses but also triggers autophagy. Autophagy is a homeostatic process that exerts multiple effects on innate immunity. However, systematic evidence linking the cGAS–STING pathway and autophagy is still lacking. Therefore, one goal of this review is to summarize the known mechanisms of autophagy induced by the cGAS–STING pathway and their consequences. The cGAS–STING pathway can trigger canonical autophagy through liquid-phase separation of the cGAS–DNA complex, interaction of cGAS and Beclin-1, and STING-triggered ER stress–mTOR signaling. Furthermore, both cGAS and STING can induce non-canonical autophagy via LC3-interacting regions and binding with LC3. Subsequently, autophagy induced by the cGAS–STING pathway plays crucial roles in balancing innate immune responses, maintaining intracellular environmental homeostasis, alleviating liver injury, and limiting tumor growth and transformation.

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An Update on Innate Immune Responses during SARS-CoV-2 Infection

Viruses ◽

10.3390/v13102060 ◽

2021 ◽

Vol 13 (10) ◽

pp. 2060

Author(s):

Yu Zhang ◽

Shuaiyin Chen ◽

Yuefei Jin ◽

Wangquan Ji ◽

Weiguo Zhang ◽

...

Keyword(s):

Immune Responses ◽

Innate Immune ◽

Type I Interferons ◽

Organ Injury ◽

Type I ◽

Innate Immune Responses ◽

Innate Immune Signaling ◽

Viral Proteases ◽

Viral Components ◽

The Relationship

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a member of the Coronaviridae family, which is responsible for the COVID-19 pandemic followed by unprecedented global societal and economic disruptive impact. The innate immune system is the body’s first line of defense against invading pathogens and is induced by a variety of cellular receptors that sense viral components. However, various strategies are exploited by SARS-CoV-2 to disrupt the antiviral innate immune responses. Innate immune dysfunction is characterized by the weak generation of type I interferons (IFNs) and the hypersecretion of pro-inflammatory cytokines, leading to mortality and organ injury in patients with COVID-19. This review summarizes the existing understanding of the mutual effects between SARS-CoV-2 and the type I IFN (IFN-α/β) responses, emphasizing the relationship between host innate immune signaling and viral proteases with an insight on tackling potential therapeutic targets.

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Role of Type I Interferons on Filovirus Pathogenesis

Vaccines ◽

10.3390/vaccines7010022 ◽

2019 ◽

Vol 7 (1) ◽

pp. 22 ◽

Cited By ~ 1

Author(s):

Beatriz Escudero-Pérez ◽

César Muñoz-Fontela

Keyword(s):

Animal Models ◽

Immune Responses ◽

Marburg Virus ◽

Type I Interferons ◽

Type I ◽

Adaptive Immune ◽

Antigen Presenting

Filoviruses, such as Ebola and Marburg virus, encode viral proteins with the ability to counteract the type I interferon (IFN-I) response. These IFN-I antagonist proteins are crucial to ensure virus replication, prevent an antiviral state in infected and bystander cells, and impair the ability of antigen-presenting cells to initiate adaptive immune responses. However, in recent years, a number of studies have underscored the conflicting data between in vitro studies and in vivo data obtained in animal models and clinical studies during outbreaks. This review aims to summarize these data and to discuss the relative contributions of IFN-α and IFN-β to filovirus pathogenesis in animal models and humans. Finally, we evaluate the putative utilization of IFN-I in post-exposure therapy and its implications as a biomarker of vaccine efficacy.

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