scholarly journals Protein Kinase R Restricts the Intracellular Survival of Mycobacterium tuberculosis by Promoting Selective Autophagy

2021 ◽  
Vol 11 ◽  
Author(s):  
Robin Smyth ◽  
Stefania Berton ◽  
Nusrah Rajabalee ◽  
Therese Chan ◽  
Jim Sun
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Protein Kinase ◽  
Intracellular Survival ◽  
Signaling Proteins ◽  
Bafilomycin A1 ◽  
Intracellular Replication ◽  
Protein Kinase R ◽  
Selective Autophagy ◽  
Cellular Antiviral Response ◽  
Host Target

Tuberculosis (TB) is a deadly infectious lung disease caused by the pathogenic bacterium Mycobacterium tuberculosis (Mtb). The identification of macrophage signaling proteins exploited by Mtb during infection will enable the development of alternative host-directed therapies (HDT) for TB. HDT strategies will boost host immunity to restrict the intracellular replication of Mtb and therefore hold promise to overcome antimicrobial resistance, a growing crisis in TB therapy. Protein Kinase R (PKR) is a key host sensor that functions in the cellular antiviral response. However, its role in defense against intracellular bacterial pathogens is not clearly defined. Herein, we demonstrate that expression and activation of PKR is upregulated in macrophages infected with Mtb. Immunological profiling of human THP-1 macrophages that overexpress PKR (THP-PKR) showed increased production of IP-10 and reduced production of IL-6, two cytokines that are reported to activate and inhibit IFNγ-dependent autophagy, respectively. Indeed, sustained expression and activation of PKR reduced the intracellular survival of Mtb, an effect that could be enhanced by IFNγ treatment. We further demonstrate that the enhanced anti-mycobacterial activity of THP-PKR macrophages is mediated by a mechanism dependent on selective autophagy, as indicated by increased levels of LC3B-II that colocalize with intracellular Mtb. Consistent with this mechanism, inhibition of autophagolysosome maturation with bafilomycin A1 abrogated the ability of THP-PKR macrophages to limit replication of Mtb, whereas pharmacological activation of autophagy enhanced the anti-mycobacterial effect of PKR overexpression. As such, PKR represents a novel and attractive host target for development of HDT for TB, and our data suggest value in the design of more specific and potent activators of PKR.

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 Immunoproteasome induction is suppressed in hepatitis C virus-infected cells in a protein kinase R-dependent manner

2016 ◽  
Vol 48 (11) ◽  
pp. e270-e270 ◽  
Author(s):  
In Soo Oh ◽  
Kathrin Textoris-Taube ◽  
Pil Soo Sung ◽  
Wonseok Kang ◽  
Xenia Gorny ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Protein Kinase ◽  
Dependent Manner ◽  
Protein Kinase R ◽  
Infected Cells

scholarly journals Tilapia dsRNA-activated protein kinase R (PKR): An interferon-induced antiviral effector with translation inhibition activity

2021 ◽  
Vol 112 ◽  
pp. 74-80
Author(s):  
Zhen Gan ◽  
Jun Cheng ◽  
Jing Hou ◽  
Shannan Chen ◽  
Hongli Xia ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Inhibition Activity ◽  
Protein Kinase R ◽  
Translation Inhibition

scholarly journals Puumala and Andes Orthohantaviruses Cause Transient Protein Kinase R-Dependent Formation of Stress Granules

2019 ◽  
Vol 94 (3) ◽  
Author(s):  
Wanda Christ ◽  
Janne Tynell ◽  
Jonas Klingström
Keyword(s):  
Protein Kinase ◽  
Host Cell ◽  
Stress Responses ◽  
Stress Granules ◽  
Cell Stress ◽  
Stress Granule ◽  
Hantavirus Infection ◽  
Stress Signaling ◽  
Protein Kinase R ◽  
Granule Formation

ABSTRACT Virus infection frequently triggers host cell stress signaling resulting in translational arrest; as a consequence, many viruses employ means to modulate the host stress response. Hantaviruses are negative-sense, single-stranded RNA viruses known to inhibit host innate immune responses and apoptosis, but their impact on host cell stress signaling remains largely unknown. In this study, we investigated activation of host cell stress responses during hantavirus infection. We show that hantavirus infection causes transient formation of stress granules (SGs) but does so in only a limited proportion of infected cells. Our data indicate some cell type-specific and hantavirus species-specific variability in SG prevalence and show SG formation to be dependent on the activation of protein kinase R (PKR). Hantavirus infection inhibited PKR-dependent SG formation, which could account for the transient nature and low prevalence of SG formation observed during hantavirus infection. In addition, we report only limited colocalization of hantaviral proteins or RNA with SGs and show evidence indicating hantavirus-mediated inhibition of PKR-like endoplasmic reticulum (ER) kinase (PERK). IMPORTANCE Our work presents the first report on stress granule formation during hantavirus infection. We show that hantavirus infection actively inhibits stress granule formation, thereby escaping the detrimental effects on global translation imposed by host stress signaling. Our results highlight a previously uncharacterized aspect of hantavirus-host interactions with possible implications for how hantaviruses are able to cause persistent infection in natural hosts and for pathogenesis.

Distinct roles of protein kinase R and toll-like receptor 3 in the activation of astrocytes by viral stimuli

Glia ◽  
2006 ◽  
Vol 55 (3) ◽  
pp. 239-252 ◽  
Author(s):  
Pamela A. Carpentier ◽  
Bryan R. Williams ◽  
Stephen D. Miller
Keyword(s):  
Protein Kinase ◽  
Toll Like Receptor ◽  
Protein Kinase R
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