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Three distinct pathways for double‐strand break formation at palindromic repeats in yeast (735.10)
Author(s) -
Sheng Ziwei,
Saini Natalie,
Haber James,
Lobachev Kirill
Publication year - 2014
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.28.1_supplement.735.10
Subject(s) - palindrome , palindromic sequence , biology , schizosaccharomyces pombe , genetics , dna , saccharomyces cerevisiae , microbiology and biotechnology , gene , crispr
Palindromic DNA sequences capable of forming cruciform and hairpin structures are strong hot‐spots for chromosomal rearrangements in a variety of organisms including humans. Previously, we demonstrated that palindromes induce chromosomal instability by triggering double‐strand breaks (DSBs) in yeast. In this study, we uncovered three mechanisms for DSB formation at palindromic loci. In E. coli and S. pombe, the SbcCD and Rad23‐Rad50‐Nbs1 complex were implicated in metabolism of palindrome during replication. On the other hand, the S. cerevisiae homolog Mre11/Rad50/Xrs2 (MRX) complex does not induce DSBs at the Alu quasi‐palindrome. We found that the perfect palindromes, compared to quasi‐palindrome, create a more prominent replication barrier and at the same time, are subject to MRX attack. Mus81‐Mms4 generates breakage in mrx mutants at palindromes formed by two URA3 genes but not at other perfect palindromes. The fact that hairpin‐capped DSB formation is not eliminated in mrx or Δmus81 mutant implies the existence of an alternative pathway for chromosomal breakage at palindromic loci. We developed a system that allows creating a quasi‐palindrome in a controllable manner throughout the cell cycle. The advantage of this system is that the pre‐existing population of DSBs is absent therefore allowing the visualization of DSBs in each cell cycle stage. Surprisingly, we found that, break formation is tightly regulated and occurs mainly in the G2 stage. We are using this system to examine the roles of cell cycle kinases and the other known structure‐specific nucleases in the generation of hairpin‐capped breaks.