Send to

Choose Destination
J Biol Chem. 2019 Oct 11;294(41):15037-15051. doi: 10.1074/jbc.RA119.008053. Epub 2019 Aug 19.

Genome-wide mapping of regions preferentially targeted by the human DNA-cytosine deaminase APOBEC3A using uracil-DNA pulldown and sequencing.

Author information

Department of Chemistry, Wayne State University, Detroit, Michigan 48202.
Center for Molecular Medicine and Genetics, Wayne State University, Wayne State University School of Medicine, Detroit, Michigan 48201.
Department of Pathology and Cancer Center, Massachusetts General Hospital, Boston, Massachusetts 02114.
Department of Chemistry, Wayne State University, Detroit, Michigan 48202
Department of Biochemistry, Microbiology and Immunology, Wayne State University School of Medicine, Detroit, Michigan 48201.


Activation-induced deaminase (AID) and apolipoprotein B mRNA-editing enzyme catalytic subunit (APOBEC) enzymes convert cytosines to uracils, creating signature mutations that have been used to predict sites targeted by these enzymes. Mutation-based targeting maps are distorted by the error-prone or error-free repair of these uracils and by selection pressures. To directly map uracils created by AID/APOBEC enzymes, here we used uracil-DNA glycosylase and an alkoxyamine to covalently tag and sequence uracil-containing DNA fragments (UPD-Seq). We applied this technique to the genome of repair-defective, APOBEC3A-expressing bacterial cells and created a uracilation genome map, i.e. uracilome. The peak uracilated regions were in the 5'-ends of genes and operons mainly containing tRNA genes and a few protein-coding genes. We validated these findings through deep sequencing of pulldown regions and whole-genome sequencing of independent clones. The peaks were not correlated with high transcription rates or stable RNA:DNA hybrid formation. We defined the uracilation index (UI) as the frequency of occurrence of TT in UPD-Seq reads at different original TC dinucleotides. Genome-wide UI calculation confirmed that APOBEC3A modifies cytosines in the lagging-strand template during replication and in short hairpin loops. APOBEC3A's preference for tRNA genes was observed previously in yeast, and an analysis of human tumor sequences revealed that in tumors with a high percentage of APOBEC3 signature mutations, the frequency of tRNA gene mutations was much higher than in the rest of the genome. These results identify multiple causes underlying selection of cytosines by APOBEC3A for deamination, and demonstrate the utility of UPD-Seq.


AID/APOBEC; ChIP-sequencing (ChIP-seq); DNA repair; DNA sequencing; UPD-Seq; base excision repair (BER); cytosine deaminase; mutagenesis; uracilome

[Available on 2020-10-11]

Supplemental Content

Full text links

Icon for HighWire
Loading ...
Support Center