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SON DNA-binding protein mediates macrophage autophagy and responses to intracellular infection.

ABSTRACT: Intracellular pathogens affect diverse host cellular defence and metabolic pathways. Here, we used infection with Francisella tularensis to identify SON DNA-binding protein as a central determinant of macrophage activities. RNAi knockdown of SON increases survival of human macrophages following F. tularensis infection or inflammasome stimulation. SON is required for macrophage autophagy, interferon response factor 3 expression, type I interferon response and inflammasome-associated readouts. SON knockdown has gene- and stimulus-specific effects on inflammatory gene expression. SON is required for accurate splicing and expression of GBF1, a key mediator of cis-Golgi structure and function. Chemical GBF1 inhibition has similar effects to SON knockdown, suggesting that SON controls macrophage functions at least in part by controlling Golgi-associated processes.

SUBMITTER: Gregory DJ

PROVIDER: S-EPMC7708400 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature

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SON DNA-binding protein mediates macrophage autophagy and responses to intracellular infection.

Gregory David J DJ DeLoid Glen M GM Salmon Sharon L SL Metzger Dennis W DW Kramnik Igor I Kobzik Lester L

FEBS letters 20200619 17

Intracellular pathogens affect diverse host cellular defence and metabolic pathways. Here, we used infection with Francisella tularensis to identify SON DNA-binding protein as a central determinant of macrophage activities. RNAi knockdown of SON increases survival of human macrophages following F. tularensis infection or inflammasome stimulation. SON is required for macrophage autophagy, interferon response factor 3 expression, type I interferon response and inflammasome-associated readouts. SON ...[more]

PMID: 32484234

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Project description:AimsProtein S-glutathionylation, the formation of a mixed disulfide between glutathione and protein thiols, is an oxidative modification that has emerged as a new signaling paradigm, potentially linking oxidative stress to chronic inflammation associated with heart disease, diabetes, cancer, lung disease, and aging. Using a novel, highly sensitive, and selective proteomic approach to identify S-glutathionylated proteins, we tested the hypothesis that monocytes and macrophages sense changes in their microenvironment and respond to metabolic stress by altering their protein thiol S-glutathionylation status.ResultsWe identified over 130 S-glutathionylated proteins, which were associated with a variety of cellular functions, including metabolism, transcription and translation, protein folding, free radical scavenging, cell motility, and cell death. Over 90% of S-glutathionylated proteins identified in metabolically stressed THP-1 monocytes were also found in hydrogen peroxide (H2O2)-treated cells, suggesting that H2O2 mediates metabolic stress-induced protein S-glutathionylation in monocytes and macrophages. We validated our findings in mouse peritoneal macrophages isolated from both healthy and dyslipidemic atherosclerotic mice and found that 52% of the S-glutathionylated proteins found in THP-1 monocytes were also identified in vivo. Changes in macrophage protein S-glutathionylation induced by dyslipidemia were sexually dimorphic.InnovationWe provide a novel mechanistic link between metabolic (and thiol oxidative) stress, macrophage dysfunction, and chronic inflammatory diseases associated with metabolic disorders.ConclusionOur data support the concept that changes in the extracellular metabolic microenvironment induce S-glutathionylation of proteins central to macrophage metabolism and a wide array of cellular signaling pathways and functions, which in turn initiate and promote functional and phenotypic changes in macrophages. Antioxid. Redox Signal. 25, 836-851.

Project description:BackgroundSarcoidosis is caused by Th1-type immune responses to unknown agents, and is linked to the infectious agent Propionibacterium acnes. Many strains of P. acnes isolated from sarcoid lesions cause intracellular infection and autophagy may contribute to the pathogenesis of sarcoidosis. We examined whether P. acnes induces autophagy.MethodsThree cell lines from macrophages (Raw264.7), mesenchymal cells (MEF), and epithelial cells (HeLa) were infected by viable or heat-killed P. acnes (clinical isolate from sarcoid lymph node) at a multiplicity of infection (MOI) of 100 or 1000 for 1 h. Extracellular bacteria were killed by washing and culturing infected cells with antibiotics. Samples were examined by colony assay, electron-microscopy, and fluorescence-microscopy with anti-LC3 and anti-LAMP1 antibodies. Autophagy-deficient (Atg5-/-) MEF cells were also used.ResultsSmall and large (≥5 μm in diameter) LC3-positive vacuoles containing few or many P. acnes cells (LC3-positive P. acnes) were frequently found in the three cell lines when infected by viable P. acnes at MOI 1000. LC3-positive large vacuoles were mostly LAMP1-positive. A few small LC3-positive/LAMP1-negative vacuoles were consistently observed in some infected cells for 24 h postinfection. The number of LC3-positive P. acnes was decreased at MOI 100 and completely abolished when heat-killed P. acnes was used. LC3-positive P. acnes was not found in autophagy-deficient Atg5-/- cells where the rate of infection was 25.3 and 17.6 times greater than that in wild-type Atg5+/+ cells at 48 h postinfection at MOI 100 and 1000, respectively. Electron-microscopic examination revealed bacterial cells surrounded mostly by a single-membrane including the large vacuoles and sometimes a double or multi-layered membrane, with occasional undigested bacterial cells in ruptured late endosomes or in the cytoplasm.ConclusionAutophagy was induced by intracellular P. acnes infection and contributed to intracellular bacterial killing as an additional host defense mechanism to endocytosis or phagocytosis.

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SON DNA-binding protein mediates macrophage autophagy and responses to intracellular infection.

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SON DNA-binding protein mediates macrophage autophagy and responses to intracellular infection.

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