Mechanism of palindrome-induced DNA double stranded breakage (DSBs) in yeast
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Chromosomes are dynamic cellular structures that carry genetic information; they combine the stability required for inheritance and the flexibility required for change. The departure from normal chromosome number and arrangement is underlying molecular feature of many cancers and hereditary diseases in humans. Gross chromosomal rearrangements (GCR), which can be caused by DNA double strand breakage (DSB), is a common type of chromosomal mutations [Inagaki et al., 2013]. Long palindromic sequences that are self-complementary DNA sequences and capable of forming non-B cruciform structures are one of the recognized breakpoint hotspots [Inagaki et al., 2013]. Genomic instability of palindromic DNA can be induced by some reactions that cleave the cruciform structure diagonally at the four-way junction, leading to frequent DNA breakage [Inagaki et al., 2013]. Currently, the enzymes that cause these DSBs in inverted repeats is still under questions even though there are studies showed multiple structure-specific nucleases can potentially target hairpin or cruciform structure [Schwartz, 2017], and this research aims to identify the enzyme by screening for mutants with lower frequency of DSB in yeast population that has been treated with mutagenic agent ethyl methanesulfonate (EMS). First, mutants exhibiting decreased levels of GCRs were identified. Then, hypo-GCR isolates were tested with Southern blotting to reveal possible low DSB mutants. These mutants will further be tested in comparison to the wild-type to identify the nuclease that causes DSB in palindromic DNA of yeast cells.