Active RNAP pre-initiation sites are highly mutated by cytidine deaminases in yeast, with AID targeting small RNA genes

Abstract

Cytidine deaminases are single stranded DNA mutators diversifying antibodies and restricting viral infection. Improper access to the genome leads to translocations and mutations in B cells and contributes to the mutation landscape in cancer, such as kataegis. It remains unclear how deaminases access double stranded genomes and whether off-target mutations favor certain loci, although transcription and opportunistic access during DNA repair are thought to play a role. In yeast, AID and the catalytic domain of APOBEC3G preferentially mutate transcriptionally active genes within narrow regions, 110 base pairs in width, fixed at RNA polymerase initiation sites. Unlike APOBEC3G, AID shows enhanced mutational preference for small RNA genes (tRNAs, snoRNAs and snRNAs) suggesting a putative role for RNA in its recruitment. We uncover the high affinity of the deaminases for the single stranded DNA exposed by initiating RNA polymerases (a DNA configuration reproduced at stalled polymerases) without a requirement for specific cofactors.In cells, genetic information is stored within molecules of DNA, which contain sequences of four ‘bases’ arranged in different orders. Replacing one of these bases with a different base results in a mutation, which can have a positive or negative influence on the cell.Mammals use a group of enzymes called cytidine deaminases to help defend themselves against harmful invaders. These enzymes work by introducing mutations into the DNA of viruses, microbes and even the mammal itself. For example, an enzyme called APOBEC3G can mutate the DNA of viruses to prevent them spreading around the body. Another enzyme, called AID, can mutate the genes that make antibodies—proteins that attack the invading microbes—in order to make new varieties of antibodies. Unfortunately, the enzymes sometimes target other genes, which can lead to cancer and other diseases.Cytidine deaminases can only access and mutate single strands of DNA, so most of the DNA in a cell is protected because it is in a two-stranded double helix. However, there are times when the two strands are separated, such as when a section of DNA is being repaired, or when it is being transcribed to produce a molecule of RNA, which is subsequently used to make a protein. It is not clear when cytidine deaminases are able to target single stranded DNA, and whether they need help from any other components.Now, Taylor et al. have studied how these enzymes access single stranded DNA when artificially introduced into yeast. These experiments showed that AID and APOBEC3G can access single stranded DNA without the help of any extra components. The enzymes target genes that are being transcribed to make RNA, with the DNA at the start of the transcription site being the most prone to mutation.In mammal cells, most genes are normally protected from the mutations introduced by cytidine deaminases, but this protection does not appear to work in many cancer cells. The next challenge will be to develop a better understanding of how this protection works, and to work out why it sometimes goes wrong.