Mitochondrial respiratory complex III sustains IL-10 production in activated macrophages and promotes tumor-mediated immune evasion
Data files
Dec 31, 2024 version files 69.01 KB
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Figure_1_raw_data.xlsx
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Figure_3_raw_data.xlsx
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Figure_4_raw_data.xlsx
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geo_mean_IL-10_calculations_Alessia_S3QEL_raw_data_figure_5.xls
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README.md
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S3QL_heatmap.R
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Abstract
The cytokine interleukin (IL)-10 limits the immune response and promotes resolution of acute and chronic inflammation. Because of its immunosuppressive effects, IL-10 upregulation is a common feature of tumor progression and metastasis. Most recently, IL-10 regulation has been shown to depend on mitochondria and redox-sensitive signals. We have found that Suppressor of site IIIQo Electron Leak (S3QEL) 1.2, a specific inhibitor of reactive oxygen species (ROS) production from mitochondrial Complex III, and Myxothiazol, a Complex III inhibitor, decrease IL-10 production from Lipopolysaccharide (LPS) activated macrophages. IL-10 downregulation is likely to be mediated by the suppression of c-Fos, which is a subunit of Activator Protein (AP)-1, a transcription factor required for IL-10 gene expression. S3QEL 1.2 impairs IL-10 production in vivo after LPS challenge and promotes the survival of mice bearing B16F10 melanoma, by lowering tumor growth. Our data identify a link between Complex III-dependent ROS generation and IL-10 production in macrophages, the specific targeting of which could have potential in the effort to boost anti-tumor immunity.
README: Mitochondrial respiratory complex III sustains IL-10 production in activated macrophages and promotes tumor-mediated immune evasion
https://doi.org/10.5061/dryad.zkh1893k1
Description of the data and file structure
Fig 1: Analysis of ROS production by Amplex Red assay (described in M&M) in Bone marrow-derived macrophages (BMDMs) pre-treated with DMSO, Myxothiazol, S3QEL 1.2 and MitoTEMPO and stimulated with LPS. Analysis of Maximal Respiration, Spare Respiratory Capacity and Complex III activity assessed by Seahorse technology in BMDMs pre-treated with DMSO, Myxothiazol, S3QEL 1.2, and stimulated with LPS. All values are from 3 independent experiments (biological replicate, indicated as 'replicate' in the Excel file on Dryad).
Fig 2: p values and counts of genes differentially expressed (downregulated) after S3QEL 1.2 and MYX pre-treated and LPS stimulated BMDMs, as obtained from bulk RNA-seq analysis. Data are from 3 mice from 1 independent experiment.
Fig 3: RT-qPCR (Fold change) and ELISA (pg/mL) experiments, analysing IL-10 expression in BMDMs pre-treated with DMSO, S3QEL 1.2 or Myxothiazol and primed with LPS. 2-3 mice were tested on 3 different weeks (3 independent times).
Fig 4: Quantitative data regarding phospho-c-Fos signal intensity in the nucleus, performed by confocal microscopy (and analysed and obtained with Imaris Software). The experiment shown in Fig.4 is representative of different independent experiments.
Fig 5: Geometric mean of fluorescent intensity (MFI) for IL-10 positive immune cells as performed by flow cytometry staining and analysis (see M&M and supplementary figure S8).
Cell Lines: BMDMs. Other samples: individual mice from in vivo experiments.
Description of the data and file structure
Data in this upload are organized based on the main figures they appear in, in the publication: Mitochondrial respiratory complex III sustains IL-10 production in activated macrophages and promotes tumor-mediated immune evasion.
Files and variables
File: S3QL_heatmap.R
Description: RNA-seq analysis of genes downregulated by S3QEL 1.2 and MYX in LPS-activated macrophages.
File: Figure_4_raw_data.xlsx
Description: Analysis of AP-1 presence in the nucleus of macrophages pre-treated with S3QEL 1.2 and MYX and stimulated with LPS.
Variables
- phospho-c-fos signal intensity
File: geo_mean_IL-10_calculations_Alessia_S3QEL_raw_data_figure_5.xls
Description: Analysis of IL-10 MFI in immune cells isolated from tumors of mice treated with PBS or S3QEL 1.2.
Variables
- IL-10 MFI
File: Figure_3_raw_data.xlsx
Description: Analysis of IL-10 expression after Complex III inhibition by S3QEL 1.2 and MYX and LPS priming.
Variables
- IL-10 mRNA and concentration of released protein.
File: Figure_1_raw_data.xlsx
Description: Analysis of ROS production, respiration and Complex III activity in macrophages after S3QEL 1.2 and MYX treatment and LPS stimulation.
Variables
- Fluorescence intensity and Oxygen Consumption Rate (OCR)
Code/software
To visualize and analyse the data contained in this data repository include GraphPad Prism v10.2.2, FlowJo v10.10 and Imaris v10.2.
Access information
Other publicly accessible locations of the data:
- N/A
Data was derived from the following sources:
- N/A
Methods
This dataset was collected after performing experiments on primary cultures of murine bone marrow-derived macrophages (BMDMs), murine B16F10 melanoma cells, human macrophages differentiated from peripheral blood mononuclear cells (PBMCs) and from in vivo experiments on mice.