m5C

5-methylcytosine (m5C) is a modified cytosine base in which a methyl group is attached to the carbon-5 position of the pyrimidine ring. This modification occurs in DNA and RNA across diverse organisms and contributes to the regulation of genetic information and RNA function. In DNA, m5C most commonly appears at cytosine residues within CpG dinucleotides in many animal genomes, although non-CpG methylation is also observed in plants and some developmental contexts.

DNA methylation patterns are established and maintained by DNA methyltransferases. In mammals, DNMT1 primarily maintains existing methylation during DNA replication, while DNMT3A and DNMT3B establish new methylation patterns. 5mC can be oxidized by TET enzymes to 5-hydroxymethylcytosine and further derivatives, which can contribute to demethylation processes. Aberrant DNA methylation is a hallmark of development and disease, with hypermethylation of promoter regions often associated with gene silencing and global hypomethylation linked to genome instability.

In RNA, m5C is found in tRNA, rRNA, and messenger RNA, among other RNAs. It is installed by RNA methyltransferases such as DNMT2 (TRDMT1), which methylates certain tRNAs, and the NSUN family (NSUN2, NSUN3, NSUN4, NSUN5), which modify various RNA substrates. m5C in RNA influences RNA stability, structure, and translation fidelity; dynamic changes in m5C have been linked to cellular stress responses and development.

Detection and study of m5C include methods for mapping DNA methylation, notably bisulfite sequencing, where unmethylated cytosines are converted to uracil while 5mC remains read as cytosine. Distinguishing 5mC from other oxidized forms requires specialized approaches (e.g., oxidative bisulfite sequencing). For RNA, approaches such as RNA bisulfite sequencing and antibody-based enrichment are used to map m5C sites. Clinically, aberrant m5C regulation is associated with cancer and other diseases, and inhibitors of DNA methyltransferases are used therapeutically in some cancers; mutations in RNA m5C writers can underlie developmental disorders.