Strand-biased cytosine deamination at the replication fork causes cytosine to thymine mutations in Escherichia coli

Proc Natl Acad Sci U S A. 2016 Feb 23;113(8):2176-81. doi: 10.1073/pnas.1522325113. Epub 2016 Feb 2.

Abstract

The rate of cytosine deamination is much higher in single-stranded DNA (ssDNA) than in double-stranded DNA, and copying the resulting uracils causes C to T mutations. To study this phenomenon, the catalytic domain of APOBEC3G (A3G-CTD), an ssDNA-specific cytosine deaminase, was expressed in an Escherichia coli strain defective in uracil repair (ung mutant), and the mutations that accumulated over thousands of generations were determined by whole-genome sequencing. C:G to T:A transitions dominated, with significantly more cytosines mutated to thymine in the lagging-strand template (LGST) than in the leading-strand template (LDST). This strand bias was present in both repair-defective and repair-proficient cells and was strongest and highly significant in cells expressing A3G-CTD. These results show that the LGST is accessible to cellular cytosine deaminating agents, explains the well-known GC skew in microbial genomes, and suggests the APOBEC3 family of mutators may target the LGST in the human genome.

Keywords: APOBEC3A; APOBEC3B; cancer genome mutations; kataegis; uracil-DNA glycosylase.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • APOBEC-3G Deaminase
  • Base Sequence
  • Cytidine Deaminase / genetics
  • Cytidine Deaminase / metabolism
  • Cytosine / metabolism
  • DNA / genetics
  • DNA / metabolism
  • DNA Repair / genetics
  • DNA Replication
  • DNA, Bacterial / genetics
  • DNA, Bacterial / metabolism
  • DNA, Single-Stranded / genetics
  • DNA, Single-Stranded / metabolism
  • Deamination
  • Escherichia coli / genetics*
  • Escherichia coli / metabolism*
  • Escherichia coli Proteins / genetics
  • Escherichia coli Proteins / metabolism
  • Genes, Bacterial
  • Humans
  • Mutation
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Thymine / metabolism
  • Uracil / metabolism
  • Uracil-DNA Glycosidase / genetics
  • Uracil-DNA Glycosidase / metabolism

Substances

  • DNA, Bacterial
  • DNA, Single-Stranded
  • Escherichia coli Proteins
  • Recombinant Proteins
  • Uracil
  • Cytosine
  • DNA
  • Uracil-DNA Glycosidase
  • APOBEC-3G Deaminase
  • APOBEC3G protein, human
  • Cytidine Deaminase
  • Thymine