Related Experiment Video
Updated: Feb 24, 2026
A Macrophage Reporter Cell Assay to Examine Toll-Like Receptor-Mediated NF-kB/AP-1 Signaling on Adsorbed Protein Layers on Polymeric Surfaces
Published on: January 7, 2020
BCG Increased Membrane Expression of TRIM59 Through the TLR2/ TLR4/IRF5 Pathway in RAW264.7 Macrophages
Zheng Jin1, Yuan Tian2, Dongmei Yan1
1Department of Immunology, College of basic Medical sciences, Jilin University, Xinmin Street 126#, Changchun, Jilin. China.
Objective:
In our previous study, we showed that Bacillus Calmette-Guerin (BCG)- activated macrophages have the ability to directly kill tumor cells. One of the main properties of these macrophages is the high expression of tripartite motif family protein 59 (TRIM59). This study was conducted to investigate the mechanism of BCG-induced TRIM59 expression on macrophages and to identify the subcellular localization of TRIM59.
Methods:
TRIM59 expression and TNF-α secretion were compared in RAW264.7 macrophage cells that were stimulated using BCG with or without Toll-like receptor 2/4 (TLR2/4)-neutralizing antibodies. Next, small interfering RNA (siRNA) was used to down-regulated interferon regulatory factor 5 (IRF5) gene expression in RAW264.7 cells. Transfected cells were stimulated with BCG, after which TRIM59 expression and TNF-α secretion were evaluated in cells pre-treated with siRNA or scramble control. After treatments, supernatants were co-cultured with MCA207, and cell viabilities were determined. Moreover, BCG-stimulated RAW264.7 cells were stained for TRIM59 and F4/80 expression.
Results:
In this study, we showed that TRIM59 was expressed on the membrane of RAW264.7 cells. After blocking TLR2/4, treatment with BCG failed to induce the expression of TRIM59, IRF5, and TNF-α on RAW264.7 cells. In addition, down-regulation of IRF5 inhibited TRIM59 and TNF-α expression.
Conclusion:
Our study showed that TRIM59 is a membrane protein, and that BCG treatment upregulated TRIM59 expression on macrophages via TLR2/4 and IRF5 pathways.
Insights
Bacillus Calmette-Guerin (BCG) upregulates tripartite motif family protein 59 (TRIM59) on macrophages. This occurs via Toll-like receptor 2/4 (TLR2/4) and interferon regulatory factor 5 (IRF5) pathways, with TRIM59 located on the cell membrane.
Area of Science:
- Immunology
- Cell Biology
Background:
- Macrophages activated by Bacillus Calmette-Guerin (BCG) exhibit direct tumor cell-killing capabilities.
- High expression of tripartite motif family protein 59 (TRIM59) is a key characteristic of these activated macrophages.
Purpose of the Study:
- To elucidate the mechanism behind BCG-induced TRIM59 expression in macrophages.
- To determine the subcellular localization of TRIM59.
Main Methods:
- Investigated TRIM59 and TNF-α expression in RAW264.7 cells stimulated with BCG, with and without TLR2/4 neutralization.
- Utilized siRNA to down-regulate IRF5 expression, followed by BCG stimulation and subsequent TRIM59/TNF-α evaluation.
- Assessed cell viability after co-culturing supernatants with MCA207 tumor cells.
- Performed immunofluorescence staining for TRIM59 and F4/80 on BCG-stimulated macrophages.
Main Results:
- TRIM59 was identified as a membrane protein on RAW264.7 cells.
- Blocking TLR2/4 inhibited BCG-induced TRIM59, IRF5, and TNF-α expression.
- Down-regulation of IRF5 significantly reduced TRIM59 and TNF-α expression.
Conclusions:
- BCG treatment upregulates TRIM59 expression on macrophages.
- The upregulation mechanism involves the TLR2/4 and IRF5 signaling pathways.
- TRIM59 is localized to the macrophage cell membrane.
More Related Videos
11:48Isolation Protocol of Mouse Monocyte-derived Dendritic Cells and Their Subsequent In Vitro Activation with Tumor Immune Complexes
Published on: May 31, 2018
08:34Legionella pneumophila Outer Membrane Vesicles: Isolation and Analysis of Their Pro-inflammatory Potential on Macrophages
Published on: February 22, 2017
Related Concept Videos
01:31Defense Against Bacterial Pathogens
Phagocytes
Phagocytes are the frontline soldiers of the immune system. They include neutrophils and macrophages. Neutrophils are the most abundant type of white blood cell and are quickly mobilized to the site of infection. Macrophages are larger cells that patrol...
01:16TGF - β Signaling Pathway