Book of Abstracts: Albany 2009
June 16-20 2009
© Adenine Press (2008)
Small molecules that enhance GAA/TTC fragility
Expansion of triplex-forming GAA/TTC repeats in the first intron of FXN gene results in Friedreich?s ataxia. Besides FXN, there are a number of other polymorphic GAA/TTC loci in the human genome where the size variations thus far have been considered to be a neutral event. Using yeast as a model system, we have previously demonstrated that expanded GAA/TTC repeats represent a threat to eukaryotic genome integrity by triggering double-strand breaks and gross chromosomal rearrangements. The fragility potential strongly depends on the length of the tract and orientation of the repeats relative to the replication origin which correlates with their propensity to adopt secondary structure and to block replication progression. The fragility is mediated by mismatch repair machinery and requires the MutSβ and endonuclease activity of MutLα.
In this study we investigate the effect of the triplex-specific small molecules on GAA-mediated fragility using the chromosomal arm loss assay. It has been shown previously that synthetic coralyne and azacyanine ligands promote and stabilize triplex DNA secondary structures and have low binding affinity to duplex DNA in vitro. We have found that in vivo, azacyanines 3, 4, and 5 but not coralyne stimulate (TTC)230 and (GAA)230- mediated arm loss in a dose dependent manner. Azacyanines at concentrations that induced fragility also inhibit cell growth. Over 60% of the yeast cells are arrested at G2/M stage of cell cycle indicative of DNA-damage activated checkpoint response. Moreover, mutants defective in DSB repair show hyper sensitivity to the azacyanines. These data indicate that azacyanines stabilize triplex DNAin vivo and this might trigger multiple DSBs during the S-phase which are sensed by the checkpoint surveillance system. We propose that these small molecules can be the basis for the development of novel antitumor drugs that act via the inhibition of the cellular proliferation. We also propose that azacyanines can be used to highlight triplex-containing regions in the human cells.
1School of Biology and Institute for Bioengineering and Bioscience