scholarly journals Mycobacterium tuberculosis Protein PE6 (Rv0335c), a Novel TLR4 Agonist, Evokes an Inflammatory Response and Modulates the Cell Death Pathways in Macrophages to Enhance Intracellular Survival

2021 ◽  
Vol 12 ◽  
Author(s):  
Neha Sharma ◽  
Mohd Shariq ◽  
Neha Quadir ◽  
Jasdeep Singh ◽  
Javaid A. Sheikh ◽  
...  
Keyword(s):  
Cell Death ◽  
Mycobacterium Tuberculosis ◽  
Proinflammatory Cytokines ◽  
Intracellular Survival ◽  
Immune Defense ◽  
Innate Immune ◽  
Bafilomycin A1 ◽  
Potent Inducer ◽  
Cell Functions ◽  
Cell Death Pathways

Mycobacterium tuberculosis (M. tb) is an intracellular pathogen that exploits moonlighting functions of its proteins to interfere with host cell functions. PE/PPE proteins utilize host inflammatory signaling and cell death pathways to promote pathogenesis. We report that M. tb PE6 protein (Rv0335c) is a secretory protein effector that interacts with innate immune toll-like receptor TLR4 on the macrophage cell surface and promotes activation of the canonical NFĸB signaling pathway to stimulate secretion of proinflammatory cytokines TNF-α, IL-12, and IL-6. Using mouse macrophage TLRs knockout cell lines, we demonstrate that PE6 induced secretion of proinflammatory cytokines dependent on TLR4 and adaptor Myd88. PE6 possesses nuclear and mitochondrial targeting sequences and displayed time-dependent differential localization into nucleus/nucleolus and mitochondria, and exhibited strong Nucleolin activation. PE6 strongly induces apoptosis via increased production of pro-apoptotic molecules Bax, Cytochrome C, and pcMyc. Mechanistic details revealed that PE6 activates Caspases 3 and 9 and induces endoplasmic reticulum-associated unfolded protein response pathways to induce apoptosis through increased production of ATF6, Chop, BIP, eIF2α, IRE1α, and Calnexin. Despite being a potent inducer of apoptosis, PE6 suppresses innate immune defense strategy autophagy by inducing inhibitory phosphorylation of autophagy initiating kinase ULK1. Inversely, PE6 induces activatory phosphorylation of autophagy master regulator MtorC1, which is reflected by lower conversion of autophagy markers LC3BI to LC3BII and increased accumulation of autophagy substrate p62 which is also dependent on innate immune receptor TLR4. The use of pharmacological agents, rapamycin and bafilomycin A1, confirms the inhibitory effect of PE6 on autophagy, evidenced by the reduced conversion of LC3BI to LC3BII and increased accumulation of p62 in the presence of rapamycin and bafilomycin A1. We also observed that PE6 binds DNA, which could have significant implications in virulence. Furthermore, our analyses reveal that PE6 efficiently binds iron to likely aid in intracellular survival. Recombinant Mycobacterium smegmatis (M. smegmatis) containing pe6 displayed robust growth in iron chelated media compared to vector alone transformed cells, which suggests a role of PE6 in iron acquisition. These findings unravel novel mechanisms exploited by PE6 protein to subdue host immunity, thereby providing insights relevant to a better understanding of host–pathogen interaction during M. tb infection.

scholarly journals Cell death and infection: A double-edged sword for host and pathogen survival

2011 ◽  
Vol 195 (6) ◽  
pp. 931-942 ◽  
Author(s):  
Hiroshi Ashida ◽  
Hitomi Mimuro ◽  
Michinaga Ogawa ◽  
Taira Kobayashi ◽  
Takahito Sanada ◽  
...  
Keyword(s):  
Cell Death ◽  
Host Cell ◽  
Defense Mechanism ◽  
Bacterial Pathogens ◽  
Immune Defense ◽  
Microbial Infection ◽  
Cell Death Pathways ◽  
New Therapeutic Approaches ◽  
Survival Pathways ◽  
Host Cell Death

Host cell death is an intrinsic immune defense mechanism in response to microbial infection. However, bacterial pathogens use many strategies to manipulate the host cell death and survival pathways to enhance their replication and survival. This manipulation is quite intricate, with pathogens often suppressing cell death to allow replication and then promoting it for dissemination. Frequently, these effects are exerted through modulation of the mitochondrial pro-death, NF-κB–dependent pro-survival, and inflammasome-dependent host cell death pathways during infection. Understanding the molecular details by which bacterial pathogens manipulate cell death pathways will provide insight into new therapeutic approaches to control infection.

scholarly journals Staphylococcus aureus Biofilm-Conditioned Medium Impairs Macrophage-Mediated Antibiofilm Immune Response by Upregulating KLF2 Expression

2019 ◽  
Vol 87 (4) ◽  
Author(s):  
Talib Alboslemy ◽  
Bing Yu ◽  
Tara Rogers ◽  
Min-Ho Kim
Keyword(s):  
Staphylococcus Aureus ◽  
Immune Response ◽  
Conditioned Medium ◽  
Innate Immune Response ◽  
Immune Defense ◽  
Innate Immune ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Content Type ◽  
Cell Functions

ABSTRACT Staphylococcus aureus infections associated with the formation of biofilms on medical implants or host tissue play a critical role in the persistence of chronic infections. One critical mechanism of biofilm infection that leads to persistent infection lies in the capacity of biofilms to evade the macrophage-mediated innate immune response. It is now increasingly apparent that microorganisms exploit the negative regulatory mechanisms of the pattern recognition receptor (PRR)-mediated inflammatory response to subvert host cell functions by using various virulence factors. However, the detailed molecular mechanism, along with the identity of a target molecule, underlying the evasion of the macrophage-mediated innate immune response against S. aureus infection associated with biofilm formation remains to be elucidated. Here, using an in vitro culture model of murine macrophage-like RAW 264.7 cells, we demonstrate that S. aureus biofilm-conditioned medium significantly attenuated the capacity for macrophage bactericidal and proinflammatory responses. Importantly, the responses were associated with attenuated activation of NF-κB and increased expression of Kruppel-like factor 2 (KLF2) in RAW 264.7 cells. Small interfering RNA (siRNA)-mediated silencing of KLF2 in RAW 264.7 cells could restore the activation of NF-κB toward the bactericidal activity and generation of proinflammatory cytokines in the presence of S. aureus biofilm-conditioned medium. Collectively, our results suggest that factors secreted from S. aureus biofilms might exploit the KLF2-dependent negative regulatory mechanism to subvert macrophage-mediated innate immune defense against S. aureus biofilms.

scholarly journals Burn the house, save the day: pyroptosis in pathogen restriction

Inflammasome ◽  
2016 ◽  
Vol 2 (1) ◽  
pp. 1-6 ◽  
Author(s):  
Dave Boucher ◽  
Kaiwen W. Chen ◽  
Kate Schroder
Keyword(s):  
Immune Response ◽  
Cell Death ◽  
Programmed Cell Death ◽  
Immune Cell ◽  
Innate Immune ◽  
Cytokine Storm ◽  
Cell Death Pathways ◽  
Multicellular Organisms ◽  
Pathogen Clearance ◽  
Inflammatory Caspases

AbstractMany programmed cell death pathways are essential for organogenesis, development, immunity and the maintenance of homeostasis in multicellular organisms. Pyroptosis, a highly proinflammatory form of cell death, is a critical innate immune response to prevent intracellular infection. Pyroptosis is induced upon the activation of proinflammatory caspases within macromolecular signalling platforms called inflammasomes. This article reviews our understanding of pyroptosis induction, the function of inflammatory caspases in pyroptosis execution, and the importance of pyroptosis for pathogen clearance. It also highlights the situations in which extensive pyroptosis may in fact be detrimental to the host, leading to immune cell depletion or cytokine storm. Current efforts to understand the beneficial and pathological roles of pyroptosis bring the promise of new approaches to fight infectious diseases.

scholarly journals Gene Delivery Using Baculovirus in Human Hematopoietic Stem and Progenitor Cells Requires Inhibition of Cellular Innate Immune Pathways

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 2940-2940
Author(s):  
Christi Salisbury-Ruf ◽  
Richard H. Smith ◽  
Fariba Chinian ◽  
Daisuke Araki ◽  
Keyvan Keyvanfar ◽  
...  
Keyword(s):  
Cell Death ◽  
Innate Immune ◽  
Gfp Expression ◽  
Hematopoietic Stem ◽  
Cell Death Pathways ◽  
Human Cd34 ◽  
Cd34 Cells ◽  
Stem And Progenitor Cells ◽  
Immune Pathways ◽  
Packaging Capacity

Abstract Homology-directed gene editing of hematopoietic stem and progenitor cells (HSPCs) has the potential to treat inherited blood disorders not amendable to CRISPR-Cas9 gene inactivation or single base editing. For many diseases, one of the major hurdles is viral delivery of large DNA templates needed for gene correction. Due to limited adeno-associated virus (AAV) packaging capacity other delivery approaches are needed. Baculovirus (BV), specifically Autographa californica multiple nucleopolyhedrovirus (AcMNPV), is a large double-stranded DNA (dsDNA) virus widely used for protein expression and AAV production. In addition, BV has been proposed as a potential therapeutic vector (Ono, Viruses 2018). BV does not replicate in mammalian cells, can deliver large quantities of DNA with virtually unlimited packaging capacity, and can express genes under the control of mammalian promoters. While capable of transducing human hepatic cells and some cell lines (Chen, Biotechnol Adv. 2011), to our knowledge BV transduction efficiency has not been tested in human CD34+ HSPCs or shown in any hematopoietic cell line. Here we show for the first time that BV can be used as a gene delivery vector for primary human CD34+ cells mobilized from healthy donors. We constructed VSV-G pseudotyped BV with a copGFP reporter flanked by 4kb homology arms (HAs) to ITGB2, a locus mutated in leukocyte adhesion deficiency type I (LAD-1) (Fig. A, top). As measured by qPCR, viral DNA was detected in CD34+ cells after transduction at a multiplicity of infection (MOI) of 50, suggesting vector binding and entry in these cells. However, although toxicity was not observed, GFP expression as assessed by flow cytometry was mostly undetectable (less than 0.1%). In contrast, robust (>70%) GFP expression was measured in 293A cells using the same BV vector, suggesting that an inhibitory cellular process was uniquely triggered in primary CD34+ cells following transduction with BV. Recent work has shown that BV can activate cellular innate immune pathways including toll-like receptors (TLRs) (Abe, J Virol 2009) and cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) (Amalfi S, JVI 2020) resulting in viral clearance and attenuation of gene expression (Ono, JVI 2014). We hypothesized that inhibition of activated cellular innate immune pathways may allow for more efficient BV gene expression in human CD34+ HSPCs. To examine this possibility, we tested over a dozen small molecule inhibitors at multiple doses targeting the major dsDNA sensing innate immune pathways including cGAS-STING (Fig. A, bottom). We found that a 45-minute pre-treatment with the STING inhibitor, H-151, while slightly toxic, enhanced GFP expression several fold, from less than 0.1% to an average of 1.5% in multiple independent donors (Fig. B-C). To improve viability, we also targeted cell death pathways. We tested the pan-caspase inhibitor, zVAD-FMK, which can inhibit both innate activation of gasdermin D (GSDMD), a major dsDNA sensing pathway, as well as apoptotic cell death. We additionally tested the necroptosis inhibitor Nec-1, as necroptosis can be activated in settings of apoptotic inhibition and inflammation. Notably, the combination of both inhibitors with H-151 improved not only cell viability, but also substantially enhanced GFP expression (8%), suggesting a synergistic benefit by inhibiting both innate immune activation and cell death pathways (Fig. D-E). To assess whether BV can efficiently transduce HSPCs with long-term repopulating activity, we pre-stimulated CD34+ cells for 48 hours in culture followed by transduction with BV at an MOI of 25 with our optimized drug cocktail. We examined GFP positivity in both CD34+CD38+ progenitors and CD34+CD38- HSC enriched populations by flow cytometry. After 24 hours, we found an average of 28% GFP+ CD34+CD38- cells and 8% GFP+ CD34+CD38+ progenitors (Fig. F-G). These data suggest that using our optimized approach, BV can target more primitive HSPCs. Collectively, our results lay the groundwork for future studies characterizing innate immune responses to dsDNA viruses in CD34+ cells, and highlight the potential use of BV as a delivery system for homology-directed gene editing in HSPCs. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

scholarly journals IFN-γ mediates Paneth cell death via suppression of mTOR

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Alessandra Araujo ◽  
Alexandra Safronova ◽  
Elise Burger ◽  
Américo López-Yglesias ◽  
Shilpi Giri ◽  
...  
Keyword(s):  
Cell Death ◽  
Intestinal Inflammation ◽  
Paneth Cell ◽  
Paneth Cells ◽  
Cell Integrity ◽  
Host Protection ◽  
Cell Functions ◽  
Cell Death Pathways ◽  
Inflammatory Conditions ◽  
Ifn Γ

Paneth cells constitutively produce antimicrobial peptides and growth factors that allow for intestinal homeostasis, host protection and intestinal stem cell replication. Paneth cells rely heavily on the glycolytic metabolic program, which is in part controlled by the kinase complex Mechanistic target of rapamycin (mTORC1). Yet, little is known about mTOR importance in Paneth cell integrity under steady state and inflammatory conditions. Our results demonstrate that IFN-γ, a crucial mediator of the intestinal inflammation, acts directly on murine Paneth cells to alter their mitochondrial integrity and membrane potential, resulting in an mTORC1-dependent cell death mechanism distinct from canonical cell death pathways including apoptosis, necroptosis, and pyroptosis. These results were established with the purified cytokine and a physiologically relevant common Th1-inducing human parasite Toxoplasma gondii. Given the crucial role for IFN-γ, which is a cytokine frequently associated with the development of inflammatory bowel disease (IBD) and compromised Paneth cell functions, the identified mechanisms underlying mTORC1-dependent Paneth cell death downstream of IFN-γ may provide promising novel approaches for treating intestinal inflammation.

scholarly journals Mycobacterium tuberculosis evasion of Guanylate Binding Protein-mediated host defense in mice requires the ESX1 secretion system

2020 ◽  
Author(s):  
Andrew J. Olive ◽  
Clare M. Smith ◽  
Christina E. Baer ◽  
Jörn Coers ◽  
Christopher M. Sassetti
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Host Defense ◽  
Genetic Interaction ◽  
Mycobacterial Infection ◽  
Secretion System ◽  
Immune Defense ◽  
Chromosome 3 ◽  
Intracellular Pathogens ◽  
Potent Inducer ◽  
Defense Proteins

AbstractCell-intrinsic immune mechanisms control intracellular pathogens that infect eukaryotes. The intracellular pathogen Mycobacterium tuberculosis (Mtb) evolved to withstand cell-autonomous immunity to cause persistent infections and disease. A potent inducer of cell-autonomous immunity is the lymphocyte-derived cytokine IFNγ. While the production of IFNγ by T cells is essential to protect against Mtb, it is not capable of fully eradicating Mtb infection. This suggests that Mtb evades a subset of IFNγ-mediated antimicrobial responses, yet what mechanisms Mtb resists remains unclear. The IFNγ-inducible Guanylate binding proteins (GBPs) are key host defense proteins able to control infections with intracellular pathogens. GBPs were previously shown to directly restrict Mycobacterium bovis BCG yet their role during Mtb infection has remained unknown. Here, we examine the importance of a cluster of five GBPs on mouse chromosome 3 in controlling Mycobacterial infection. While M. bovis BCG is directly restricted by GBPs, we find that the GBPs on chromosome 3 do not contribute to the control of Mtb replication or the associated host response to infection. The differential effects of GBPs during Mtb versus M. bovis BCG infection is at least partially explained by the absence of the ESX1 secretion system from M. bovis BCG, since Mtb mutants lacking the ESX1 secretion system become similarly susceptible to GBP-mediated immune defense. Therefore, this specific genetic interaction between the murine host and Mycobacteria reveals a novel function for the ESX1 virulence system in the evasion of GBP-mediated immunity.

scholarly journals Apoptosis: Implications in Viral and Mycobacterium tuberculosis infections

2017 ◽  
Vol 5 (1) ◽  
pp. 46-57
Author(s):  
Gorakh Raj Giri ◽  
Uddhav Timilsina
Keyword(s):  
Cell Death ◽  
Mycobacterium Tuberculosis ◽  
Programmed Cell Death ◽  
Host Cell ◽  
Cell Apoptosis ◽  
Bacterial Infections ◽  
Autoimmune Disorders ◽  
Immune Defense ◽  
Pathological Conditions ◽  
Host Cell Apoptosis

Apoptosis is a form of programmed cell death leading to genetically controlled self-destruction of cells. It is essential in the development, maintenance, and regulation of cells during physiological as well as pathological conditions. Deregulation of apoptotic mechanisms is associated with various pathological diseases including cancer, autoimmune disorders, viral and bacterial infections. Virus and Mycobacterium tuberculosis elicit host cell apoptosis as a part of host immune defense or pathogen dissemination. They inhibit both extrinsic and intrinsic pathways of apoptotic mechanisms facilitating pathogen survival and escape from host immune defense.Nepal Journal of Biotechnology. Dec. 2017 Vol. 5, No. 1: 46- 57

scholarly journals Suppression of Host Innate Immune Response by Hepatitis C Virus via Induction of Autophagic Degradation of TRAF6

2016 ◽  
Vol 90 (23) ◽  
pp. 10928-10935 ◽  
Author(s):  
Stephanie T. Chan ◽  
Jiyoung Lee ◽  
Mansi Narula ◽  
J.-H. James Ou
Keyword(s):  
Immune Response ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Innate Immune Response ◽  
Proinflammatory Cytokines ◽  
Host Immunity ◽  
Innate Immune ◽  
Bafilomycin A1 ◽  
Host Innate Immune Response ◽  
Autophagic Degradation

ABSTRACT Tumor necrosis factor receptor (TNFR)-associated factor 6 (TRAF6) is an important adaptor molecule that mediates the TNFR family and interleukin-1 (IL-1)/Toll-like receptor (TLR) signaling cascades. These pathways are important for the host to control viral infections. In this report, we demonstrated that hepatitis C virus (HCV) depleted TRAF6 from its host cells through a posttranslational mechanism. This depletion was independent of proteasomes, as it was not affected by the proteasome inhibitor MG132, but it was suppressed by bafilomycin A1, which led to the association of TRAF6 with autophagosomes. As bafilomycin A1 is a vacuolar ATPase inhibitor that inhibits autophagic protein degradation, these results suggested that HCV depleted TRAF6 via autophagy. The degradation of TRAF6 was apparently mediated by the p62 sequestosome protein, which is a factor important for selective autophagy, as it could bind to TRAF6 and its silencing stabilized TRAF6. The depletion of TRAF6 suppressed activation of NF-κB and induction of proinflammatory cytokines and enhanced HCV replication. In contrast, the overexpression of TRAF6 suppressed HCV replication. These results revealed a novel mechanism that was used by HCV to disrupt the host innate immune responses for viral replication and persistence. IMPORTANCE HCV can cause severe liver diseases and is one of the most important human pathogens. It establishes chronic infections in the great majority of patients that it infects, indicating that it has evolved sophisticated mechanisms to evade host immunity. TRAF6 is an important signaling molecule that mediates activation of NF-κB and expression of proinflammatory cytokines and interferons. In this study, we found that HCV infection suppressed the host innate immune response through the induction of autophagic degradation of TRAF6. This finding provided important information for further understanding how HCV evades host immunity to establish persistence.

scholarly journals Macrophage exposures to crystalline silica promote growth of Mycobacterium tuberculosis by deregulating cell death pathways.

2013 ◽  
Vol 4 ◽  
Author(s):  
Ch�vez-Gal�n Leslie ◽  
Torre-Bouscoulet Luis ◽  
P�rez-Padilla Rogelio ◽  
Ram�rez-Venegas Alejandra ◽  
Sada-Ovalle Isabel
Keyword(s):  
Cell Death ◽  
Mycobacterium Tuberculosis ◽  
Crystalline Silica ◽  
Cell Death Pathways
Sign in / Sign up

Export Citation Format

Share Document