Putative DNA modification methylase DR_C0020 of Deinococcus radiodurans is an atypical SAM dependent C-5 cytosine DNA methylase
Abstract Background Control of cellular processes by epigenetic modification of cytosine in DNA is widespread among living organisms, but, is hitherto unknown in the extremely radioresistant microbe D. radiodurans . Methods C-5 methyl cytosines (m 5 C) were detected by immuno-blotting with m 5 C-specific antibody. Site of cytosine methylation by DR_C0020 encoded protein was investigated by bisulfite sequencing. The DR_C0020 knockout mutant (Δ dcm ), constructed by site directed mutagenesis, was assessed for effect on growth, radiation resistance and proteome. Proteins were identified by mass spectrometry. Results Methylated cytosines were detected in the D. radiodurans genome. The DR_C0020 encoded protein (Dcm, NCBI accession: WP_034351354.1), whose amino acid sequence resembles m 4 C methylases, was shown to be the lone SAM-dependent C-5 cytosine methyltransferase. Purified Dcm protein was found to methylate CpN sequence with a preference for methylation of two consecutive cytosines. The Δ dcm strain completely lost m 5 C modification from its genome, had no effect on growth but became radiation sensitive. The Δ dcm cells exhibited minor alterations in the abundance of several proteins involved primarily in protein homeostasis, oxidative stress defense, metabolism, etc. Conclusion DR_C0020 encoded SAM-dependent methyltransferase Dcm is solely responsible for C-5cytosine methylation at CpN sites in the genome of D. radiodurans and regulates protein homeostasis under normal growth conditions. The protein is an unusual case of an amino methyltransferase that has evolved to producing m 5 C. General significance Although, dispensable under optimal growth conditions, the presence of m 5 C may be important for recognition of parent strand and, thus, could contribute to the extraordinary DNA repair in D. radiodurans . Highlights The genome of D. radiodurans was shown to possess C-5 methylated cytosines. DR_C0020 encoded protein (Dcm) is a SAM-dependent C-5 cytosine methyltransferase. Dcm methylates CpN sites, quite atypical for bacterial methyltransferase. DR_C0020 gene deletion leads to complete loss of m 5 C modification in the genome. Cytosine methylation is dispensable under optimal growth conditions.
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