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Updated: Nov 13, 2025
Immunostaining for DNA Modifications: Computational Analysis of Confocal Images
Published on: September 7, 2017
m6A methylation potentiates cytosolic dsDNA recognition in a sequence-specific manner
Melania Balzarolo1,2,3, Sander Engels1,2,3,4, Anja J de Jong1,2,3
1Sanquin Research, University of Amsterdam, Amsterdam, The Netherlands.
Abstract:
Nucleic acid sensing through pattern recognition receptors is critical for immune recognition of microbial infections. Microbial DNA is frequently methylated at the N6 position of adenines (m6A), a modification that is rare in mammalian host DNA. We show here how that m6A methylation of 5'-GATC-3' motifs augments the immunogenicity of synthetic double-stranded (ds)DNA in murine macrophages and dendritic cells. Transfection with m6A-methylated DNA increased the expression of the activation markers CD69 and CD86, and of Ifnβ, iNos and Cxcl10 mRNA. Similar to unmethylated cytosolic dsDNA, recognition of m6A DNA occurs independently of TLR and RIG-I signalling, but requires the two key mediators of cytosolic DNA sensing, STING and cGAS. Intriguingly, the response to m6A DNA is sequence-specific. m6A is immunostimulatory in some motifs, but immunosuppressive in others, a feature that is conserved between mouse and human macrophages. In conclusion, epigenetic alterations of DNA depend on the context of the sequence and are differentially perceived by innate cells, a feature that could potentially be used for the design of immune-modulating therapeutics.
Insights
DNA methylation (m6A) can enhance immune responses to microbial DNA, activating innate immune cells. This epigenetic modification
Area of Science:
- Immunology
- Molecular Biology
- Epigenetics
Background:
- Pattern recognition receptors (PRRs) are crucial for detecting microbial infections.
- Microbial DNA often contains N6-methyladenine (m6A) modifications, which are rare in mammalian DNA.
Purpose of the Study:
- To investigate the impact of m6A DNA methylation on the immunogenicity of synthetic DNA.
- To determine the mechanisms by which m6A DNA is recognized by innate immune cells.
Main Methods:
- Transfection of murine macrophages and dendritic cells with m6A-methylated and unmethylated DNA.
- Analysis of immune cell activation markers (CD69, CD86) and gene expression (Ifnβ, iNos, Cxcl10).
- Assessment of signaling pathways involving TLR, RIG-I, STING, and cGAS.
Main Results:
- m6A methylation of 5'-GATC-3' motifs augmented DNA immunogenicity in murine immune cells.
- Recognition of m6A DNA required STING and cGAS, independent of TLR and RIG-I.
- The immune response to m6A DNA was sequence-specific, with some motifs being immunostimulatory and others immunosuppressive.
- This sequence-specific response was conserved in human macrophages.
Conclusions:
- Epigenetic DNA modifications are context-dependent and differentially recognized by innate immune cells.
- m6A DNA methylation can modulate immune responses, offering potential for immune-modulating therapeutics.
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