The BH3-only protein Bik/Blk/Nbk inhibits nuclear translocation of activated ERK1/2 to mediate IFNγ-induced cell death

IFNγ induces cell death in epithelial cells, but the mediator for this death pathway has not been identified. In this study, we find that expression of Bik/Blk/Nbk is increased in human airway epithelial cells (AECs [HAECs]) in response to IFNγ. Expression of Bik but not mutant BikL61G induces and loss of Bik suppresses IFNγ-induced cell death in HAECs. IFNγ treatment and Bik expression increase cathepsin B and D messenger RNA levels and reduce levels of phospho–extracellular regulated kinase 1/2 (ERK1/2) in the nuclei of bik+/+ compared with bik−/− murine AECs. Bik but not BikL61G interacts with and suppresses nuclear translocation of phospho-ERK1/2, and suppression of ERK1/2 activation inhibits IFNγ- and Bik-induced cell death. Furthermore, after prolonged exposure to allergen, hyperplastic epithelial cells persist longer, and nuclear phospho-ERK is more prevalent in airways of IFNγ−/− or bik−/− compared with wild-type mice. These results demonstrate that IFNγ requires Bik to suppress nuclear localization of phospho-ERK1/2 to channel cell death in AECs.

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Pim1 kinase protects airway epithelial cells from cigarette smoke-induced damage and airway inflammation

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00156.2013 ◽

2014 ◽

Vol 307 (3) ◽

pp. L240-L251 ◽

Cited By ~ 19

Author(s):

M. de Vries ◽

I. H. Heijink ◽

R. Gras ◽

L. E. den Boef ◽

M. Reinders-Luinge ◽

...

Keyword(s):

Cell Death ◽

Membrane Potential ◽

Epithelial Cells ◽

Airway Inflammation ◽

Mitochondrial Membrane Potential ◽

Mitochondrial Membrane ◽

Airway Epithelial Cells ◽

Airway Epithelial ◽

Wild Type ◽

Deficient Mice

Exposure to cigarette smoke (CS) is the main risk factor for developing chronic obstructive pulmonary disease and can induce airway epithelial cell damage, innate immune responses, and airway inflammation. We hypothesized that cell survival factors might decrease the sensitivity of airway epithelial cells to CS-induced damage, thereby protecting the airways against inflammation upon CS exposure. Here, we tested whether Pim survival kinases could protect from CS-induced inflammation. We determined expression of Pim kinases in lung tissue, airway inflammation, and levels of keratinocyte-derived cytokine (KC) and several damage-associated molecular patterns in bronchoalveolar lavage in mice exposed to CS or air. Human bronchial epithelial BEAS-2B cells were treated with CS extract (CSE) in the presence or absence of Pim1 inhibitor and assessed for loss of mitochondrial membrane potential, induction of cell death, and release of heat shock protein 70 (HSP70). We observed increased expression of Pim1, but not of Pim2 and Pim3, in lung tissue after exposure to CS. Pim1-deficient mice displayed a strongly enhanced neutrophilic airway inflammation upon CS exposure compared with wild-type controls. Inhibition of Pim1 activity in BEAS-2B cells increased the loss of mitochondrial membrane potential and reduced cell viability upon CSE treatment, whereas release of HSP70 was enhanced. Interestingly, we observed release of S100A8 but not of double-strand DNA or HSP70 in Pim1-deficient mice compared with wild-type controls upon CS exposure. In conclusion, we show that expression of Pim1 protects against CS-induced cell death in vitro and neutrophilic airway inflammation in vivo. Our data suggest that the underlying mechanism involves CS-induced release of S100A8 and KC.

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The uptake of a Klebsiella pneumoniae capsule polysaccharide mutant triggers an inflammatory response by human airway epithelial cells

Microbiology ◽

10.1099/mic.0.28285-0 ◽

2006 ◽

Vol 152 (2) ◽

pp. 555-566 ◽

Cited By ~ 55

Author(s):

Verónica Regueiro ◽

Miguel A. Campos ◽

Jaume Pons ◽

Sebastián Albertí ◽

José A. Bengoechea

Keyword(s):

Epithelial Cells ◽

Klebsiella Pneumoniae ◽

Inflammatory Response ◽

Nuclear Translocation ◽

A549 Cells ◽

Airway Epithelial Cells ◽

Airway Epithelial ◽

Wild Type ◽

Cellular Activation ◽

Epithelial Cell Surface

The means by which airway epithelial cells sense a bacterial infection and which intracellular signalling pathways are activated upon infection are poorly understood. A549 cells and human primary airway cells (NHBE) were used to investigate the response to infection with Klebsiella pneumoniae. Infection of A549 and NHBE with K. pneumoniae 52K10, a capsule polysaccharide (CPS) mutant, increased the surface levels of ICAM-1 and caused the release of IL-8. By contrast, the wild-type strain did not elicit these responses. Consistent with a functional role for these responses, there was a correlation between ICAM-1 levels and the number of adherent leukocytes on the epithelial cell surface. In addition, treatment of neutrophils with IL-8 enhanced their ability to kill K. pneumoniae. Strain 52K10 was internalized by A549 cells more efficiently than the wild-type, and when infections with 52K10 were performed in the presence of cytochalasin D the inflammatory response was abrogated. These findings suggest that cellular activation is mediated by bacterial internalization and that CPS prevents the activation through the blockage of bacterial adhesion and uptake. Collectively, the results indicate that bacterial internalization by airway epithelial cells could be the triggering signal for the activation of the innate immune system of the airway. Infection of A549 cells by 52K10 was shown to trigger the nuclear translocation of NF-κB. Evidence is presented showing that 52K10 activated IL-8 production through Toll-like receptor (TLR) 2 and TLR4 pathways and that A549 cells could use soluble CD14 as TLR co-receptor.

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Deacetylation of p53 induces autophagy by suppressing Bmf expression

The Journal of Cell Biology ◽

10.1083/jcb.201205064 ◽

2013 ◽

Vol 201 (3) ◽

pp. 427-437 ◽

Cited By ~ 30

Author(s):

Amelia U. Contreras ◽

Yohannes Mebratu ◽

Monica Delgado ◽

Gilbert Montano ◽

Chien-an A. Hu ◽

...

Keyword(s):

Cell Death ◽

Messenger Rna ◽

Histone Deacetylase Inhibitors ◽

Interferon Γ ◽

Airway Epithelial Cells ◽

Dependent Manner ◽

Airway Epithelial ◽

Independent Manner ◽

Rich Domain ◽

Ifn Γ

Interferon γ (IFN-γ)–induced cell death is mediated by the BH3-only domain protein, Bik, in a p53-independent manner. However, the effect of IFN-γ on p53 and how this affects autophagy have not been reported. The present study demonstrates that IFN-γ down-regulated expression of the BH3 domain-only protein, Bmf, in human and mouse airway epithelial cells in a p53-dependent manner. p53 also suppressed Bmf expression in response to other cell death–stimulating agents, including ultraviolet radiation and histone deacetylase inhibitors. IFN-γ did not affect Bmf messenger RNA half-life but increased nuclear p53 levels and the interaction of p53 with the Bmf promoter. IFN-γ–induced interaction of HDAC1 and p53 resulted in the deacetylation of p53 and suppression of Bmf expression independent of p53’s proline-rich domain. Suppression of Bmf facilitated IFN-γ–induced autophagy by reducing the interaction of Beclin-1 and Bcl-2. Furthermore, autophagy was prominent in cultured bmf−/− but not in bmf+/+ cells. Collectively, these observations show that deacetylation of p53 suppresses Bmf expression and facilitates autophagy.

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A Type IV Secretion System Contributes to Intracellular Survival and Replication of Burkholderia cenocepacia

Infection and Immunity ◽

10.1128/iai.00451-08 ◽

2008 ◽

Vol 76 (12) ◽

pp. 5447-5455 ◽

Cited By ~ 28

Author(s):

S. Umadevi Sajjan ◽

Lisa A. Carmody ◽

Carlos F. Gonzalez ◽

John J. LiPuma

Keyword(s):

Epithelial Cells ◽

Airway Epithelial Cells ◽

Secretion System ◽

Type Iv Secretion ◽

Endocytic Pathway ◽

Burkholderia Cenocepacia ◽

Type Iv Secretion System ◽

Airway Epithelial ◽

Wild Type ◽

Type Iv

ABSTRACT Burkholderia cenocepacia is an important respiratory pathogen in persons with cystic fibrosis (CF). Recent studies indicate that B. cenocepacia survives within macrophages and airway epithelial cells in vitro by evading endosome-lysosome fusion. We investigated the role of a plasmid-encoded type IV secretion system in the intracellular survival, replication, and processing of B. cenocepacia. Both a wild-type strain (K56-2) and its type IV secretion system mutant (designated LC101) entered and replicated in CF airway epithelial cells and monocyte-derived macrophages. However, significantly more intracellular K56-2 than LC101 bacteria were found in both cell types at 24 h postinfection. Colocalization of bacteria with markers of the classical endocytic pathway indicated that although both K56-2 and LC101 reside transiently in early endosomes, a greater proportion of the mutant bacteria are targeted to lysosomal degradation. In contrast, wild-type bacteria escape from the classical endocytic pathway and traffic to the endoplasmic reticulum, where they replicate. Our results show that the intracellular processing of B. cenocepacia is similar in both professional and nonprofessional phagocytes and that a functional plasmid-encoded type IV secretion system contributes to the survival and replication of B. cenocepacia in eukaryotic cells.

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Activity and inhibition of prostasin and matriptase on apical and basolateral surfaces of human airway epithelial cells

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00303.2011 ◽

2012 ◽

Vol 303 (2) ◽

pp. L97-L106 ◽

Cited By ~ 11

Author(s):

Shilpa Nimishakavi ◽

Marina Besprozvannaya ◽

Wilfred W. Raymond ◽

Charles S. Craik ◽

Dieter C. Gruenert ◽

...

Keyword(s):

Cystic Fibrosis ◽

Epithelial Cells ◽

Cell Surface ◽

Selective Inhibition ◽

Airway Epithelial Cells ◽

Bronchial Epithelial Cells ◽

Apical Surface ◽

Airway Epithelial ◽

Wild Type ◽

Bronchial Epithelial

Prostasin is a membrane-anchored protease expressed in airway epithelium, where it stimulates salt and water uptake by cleaving the epithelial Na+ channel (ENaC). Prostasin is activated by another transmembrane tryptic protease, matriptase. Because ENaC-mediated dehydration contributes to cystic fibrosis (CF), prostasin and matriptase are potential therapeutic targets, but their catalytic competence on airway epithelial surfaces has been unclear. Seeking tools for exploring sites and modulation of activity, we used recombinant prostasin and matriptase to identify substrate t-butyloxycarbonyl-l-Gln-Ala-Arg-4-nitroanilide (QAR-4NA), which allowed direct assay of proteases in living cells. Comparisons of bronchial epithelial cells (CFBE41o−) with and without functioning cystic fibrosis transmembrane conductance regulator (CFTR) revealed similar levels of apical and basolateral aprotinin-inhibitable activity. Although recombinant matriptase was more active than prostasin in hydrolyzing QAR-4NA, cell surface activity resisted matriptase-selective inhibition, suggesting that prostasin dominates. Surface biotinylation revealed similar expression of matriptase and prostasin in epithelial cells expressing wild-type vs. ΔF508-mutated CFTR. However, the ratio of mature to inactive proprostasin suggested surface enrichment of active enzyme. Although small amounts of matriptase and prostasin were shed spontaneously, prostasin anchored to the cell surface by glycosylphosphatidylinositol was the major contributor to observed QAR-4NA-hydrolyzing activity. For example, the apical surface of wild-type CFBE41o− epithelial cells express 22% of total, extractable, aprotinin-inhibitable, QAR-4NA-hydrolyzing activity and 16% of prostasin immunoreactivity. In conclusion, prostasin is present, mature and active on the apical surface of wild-type and CF bronchial epithelial cells, where it can be targeted for inhibition via the airway lumen.

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Nuclear Translocation of Lactosylated Poly-l-lysine/cDNA Complex in Cystic Fibrosis Airway Epithelial Cells

Molecular Therapy ◽

10.1006/mthe.2001.0332 ◽

2001 ◽

Vol 3 (6) ◽

pp. 831-841 ◽

Cited By ~ 47

Author(s):

Daniel T. Klink ◽

Simon Chao ◽

Mary Catherine Glick ◽

Thomas F. Scanlin

Keyword(s):

Cystic Fibrosis ◽

Epithelial Cells ◽

Nuclear Translocation ◽

Airway Epithelial Cells ◽

Airway Epithelial ◽

Cystic Fibrosis Airway

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Role of f-box factor foxj1 in differentiation of ciliated airway epithelial cells

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00170.2003 ◽

2004 ◽

Vol 286 (4) ◽

pp. L650-L657 ◽

Cited By ~ 110

Author(s):

Yingjian You ◽

Tao Huang ◽

Edward J. Richer ◽

Jens-Erik Harboe Schmidt ◽

Joseph Zabner ◽

...

Keyword(s):

Epithelial Cell ◽

Epithelial Cells ◽

Basal Body ◽

Ciliated Cell ◽

Airway Epithelial Cells ◽

Cell Phenotype ◽

Basal Bodies ◽

Airway Epithelial ◽

Ciliated Cells ◽

Wild Type

Factors required for commitment of an undifferentiated airway epithelial cell to a ciliated cell are unknown. Cell ultrastructure analysis indicates ciliated cell commitment activates a multistage program involving synthesis of cilia precursor proteins and assembly of macromolecular complexes. Foxj1 is an f-box transcription factor expressed in ciliated cells and shown to be required for cilia formation by gene deletion in a mouse model. To identify a specific role for foxj1 in directing the ciliated cell phenotype, we evaluated the capacity of foxj1 to induce ciliogenesis and direct cilia assembly. In a primary culture model of wild-type mouse airway epithelial cells, foxj1 expression preceded the appearance of cilia and in cultured foxj1 null cells cilia did not develop. Delivery of foxj1 to polarized epithelial cell lines and primary cultured alveolar epithelial cells failed to promote ciliogenesis. Similarly, delivery of foxj1 to wild-type airway epithelial cells did not enhance the total number of ciliated cells. In contrast, delivery of foxj1 to null cells resulted in the appearance of cilia. Analysis revealed that, in the absence of foxj1, null cells contained cilia precursor basal bodies, indicating prior commitment to ciliogenesis. However, the basal bodies were disorganized within the apical compartment and failed to dock with the apical membrane. Reconstitution of foxj1 in null cells restored normal basal body organization, resulting in axoneme growth. Thus foxj1 functions in late-stage ciliogenesis to regulate programs promoting basal body docking and axoneme formation in cells previously committed to the ciliated cell phenotype.

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Resveratrol induces glutathione synthesis by activation of Nrf2 and protects against cigarette smoke-mediated oxidative stress in human lung epithelial cells

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00361.2007 ◽

2008 ◽

Vol 294 (3) ◽

pp. L478-L488 ◽

Cited By ~ 267

Author(s):

Aruna Kode ◽

Saravanan Rajendrasozhan ◽

Samuel Caito ◽

Se-Ran Yang ◽

Ian L. Megson ◽

...

Keyword(s):

Oxidative Stress ◽

Epithelial Cells ◽

Posttranslational Modifications ◽

Nuclear Translocation ◽

Airway Epithelial Cells ◽

Lung Epithelial Cells ◽

Oxygen Species ◽

Airway Epithelial ◽

Alveolar Epithelial ◽

Lung Epithelial

Nuclear erythroid-related factor 2 (Nrf2), a redox-sensitive transcription factor, is involved in transcriptional regulation of many antioxidant genes, including glutamate-cysteine ligase (GCL). Cigarette smoke (CS) is known to cause oxidative stress and deplete glutathione (GSH) levels in alveolar epithelial cells. We hypothesized that resveratrol, a polyphenolic phytoalexin, has antioxidant signaling properties by inducing GSH biosynthesis via the activation of Nrf2 and protects lung epithelial cells against CS-mediated oxidative stress. Treatment of human primary small airway epithelial and human alveolar epithelial (A549) cells with CS extract (CSE) dose dependently decreased GSH levels and GCL activity, effects that were associated with enhanced production of reactive oxygen species. Resveratrol restored CSE-depleted GSH levels by upregulation of GCL via activation of Nrf2 and also quenched CSE-induced release of reactive oxygen species. Interestingly, CSE failed to induce nuclear translocation of Nrf2 in A549 and small airway epithelial cells. On the contrary, Nrf2 was localized in the cytosol of alveolar and airway epithelial cells due to CSE-mediated posttranslational modifications such as aldehyde/carbonyl adduct formation and nitration. On the other hand, resveratrol attenuated CSE-mediated Nrf2 modifications, thereby inducing its nuclear translocation associated with GCL gene transcription, as demonstrated by GCL-promoter reporter and Nrf2 small interfering RNA approaches. Thus resveratrol attenuates CSE-mediated GSH depletion by inducing GSH synthesis and protects epithelial cells by reversing CSE-induced posttranslational modifications of Nrf2. These data may have implications in dietary modulation of antioxidants in treatment of chronic obstructive pulmonary disease.

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