Active RNAP pre-initiation sites are highly mutated by cytidine deaminases in yeast, with AID targeting small RNA
Benjamin J M Taylor1, Yee Ling Wu1, Cristina Rada1
1Protein and Nucleic Acid Chemistry Division, Medical Research Council Laboratory of Molecular Biology, Cambridge, United Kingdom.
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.
Insights
Cytidine deaminases like AID bind single-stranded DNA at RNA polymerase initiation sites, explaining their mutation patterns in genes. This mechanism clarifies how these enzymes access and modify DNA, impacting antibody diversity and cancer mutations.
Area of Science:
- Molecular Biology
- Genetics
- Immunology
Background:
- Cytidine deaminases are crucial for antibody diversification and viral restriction but can cause genomic instability.
- Mechanisms of deaminase access to double-stranded DNA and off-target mutation preferences remain incompletely understood.
- Transcription and DNA repair are hypothesized roles in deaminase genome access.
Purpose of the Study:
- To investigate how cytidine deaminases access double-stranded DNA genomes.
- To determine if specific DNA configurations or loci are favored by deaminases.
- To elucidate the role of transcription and RNA polymerase in deaminase recruitment and activity.
Main Methods:
- Experiments conducted in yeast models.
- Analysis of mutation patterns for AID and APOBEC3G catalytic domain.
- Assessment of deaminase affinity for single-stranded DNA structures.
Main Results:
- AID and APOBEC3G preferentially mutate transcriptionally active genes in narrow regions (110 bp) at RNA polymerase initiation sites.
- AID exhibits a stronger preference for small RNA genes (tRNAs, snoRNAs, snRNAs), suggesting RNA involvement in its recruitment.
- Deaminases show high affinity for single-stranded DNA exposed by initiating RNA polymerases, a configuration also found at stalled polymerases.
Conclusions:
- Cytidine deaminases access DNA through single-stranded regions generated by RNA polymerase activity, not requiring specific cofactors.
- This mechanism explains the observed mutation patterns and locus specificity of AID and APOBEC3G.
- Findings provide insight into antibody gene diversification and the role of deaminases in cancer mutagenesis.
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