Abstract

Background: Replication-independent endogenous double-strand breaks (RIND-EDSBs) occur in both humans and yeast in the absence of inductive agents and DNA replication. In human cells, RIND-EDSBs are hypermethylated, preferentially retained in the heterochromatin and unbound by γ-H2AX. In single gene deletion yeast strains, the RIND-EDSB levels are altered; the number of RIND-EDSBs is higher in strains with deletions of histone deacetylase, endonucleases, topoisomerase, or DNA repair regulators, but lower in strains with deletions of the high-mobility group box proteins or Sir2. In summary, RIND-EDSBs are different from pathologic DSBs in terms of their causes and consequences. In this study, we identified the nucleotide sequences surrounding RIND-EDSBs and investigated the features of these sequences as well as their break locations. Results: In recent work, we detected RIND-EDSBs using ligation mediated PCR. In this study, we sequenced RIND-EDSB PCR products of resting state Saccharomyces cerevisiae using next-generation sequencing to analyze RIND-EDSB sequences. We found that the break locations are scattered across a number of chromosomes. The number of breaks correlated with the size of the chromosomes. Most importantly, the break occurrences had sequence pattern specificity. Specifically, the majority of the breaks occurred immediately after the sequence " ACGT" (P = 2.2E-156). Because the " ACGT" sequence does not occur primarily in the yeast genome, this specificity of the " ACGT" sequence cannot be attributed to chance. Conclusions: RIND-EDSBs occur non-randomly; that is, they are produced and retained by specific mechanisms. Because these particular mechanisms regulate their generation and they possess potentially specific functions, RIND-EDSBs could be epigenetic marks.