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bioRxiv · 10.64898/2026.10.20.760421

The relevance of GGCC methylation for cyanobacterial gene expression regulation

Stappert, N.·Schmidt, N.·Reimann, V.·Hagemann, M.·Hess, W. R.
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Abstract

Epigenetic DNA modifications are pivotal in eukaryotic gene expression, but their regulatory significance in bacteria is less understood. In the Synechocystis 6803 genome, the DNA-specific methyltransferase M.Ssp6803II modifies the first cytosine in the 38,512 GGCC motifs, forming N4-methylcytosine (GGm4CC). Previous analyses showed that deletion of the gene encoding M.Ssp6803II ({triangleup}sll0729), caused a bluish phenotype due to reduced expression of the hemJ gene. This effect was caused by the lacking GGCC methylation in the discriminator sequence of the hemJ promoter in {triangleup}sll0729. In the present study, we analyzed the impact of GGm4CC methylation on the overall gene expression. Bioinformatic analyses of multiple, independently obtained DNA microarray datasets revealed reproducible changes in the expression of more than 300 genes. We selected 26 genes that had the GGCC methylation motif in different locations of their promoter sequences and verified their differential expression through qPCR experiments. Classification of the promoters according to the relative position of GGCC motifs suggested context-specific effects on gene expression. These findings imply that GGm4CC methylation via DNA methyltransferase M.Ssp6803II has an important impact on proper gene expression and thus, cytosine N4-methylation is a relevant epigenetic marker in Synechocystis and likely many other bacteria.

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Stappert, N., Schmidt, N., Reimann, V., Hagemann, M., Hess, W. R.. 2026-10-21. The relevance of GGCC methylation for cyanobacterial gene expression regulation. https://doi.org/10.64898/2026.10.20.760421

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