A Saccharomyces cerevisiae Model for Overexpression of Ntg1 a Base Excision DNA Repair Protein Reveals Novel Genetic Interactions

Abstract

The Base Excision Repair (BER) pathway repairs oxidative DNA damage, a common and detrimental form of damage to the genome. Although biochemical steps in BER have been well define, little is understood about how the pathway is regulated. Such regulation is critical, as cells must respond rapidly to DNA damage while avoiding aberrant activation of repair proteins that can produce DNA damage as intermediates in the repair pathway. Indeed, overexpression of the human BER protein, NTHL1, a DNA N-glycosylase, can cause genomic instability and early cellular hallmarks of cancer. We developed a Saccharomyces cerevisiae model to explore how overexpression of NTHL1 may impair cellular function. Overexpression of Ntg1, the budding yeast orthologue of NTHL1, impairs cell growth. To dissect mechanisms underlying this growth defect, we overexpressed either wild-type Ntg1 or a catalytically inactive variant of Ntg1 (ntg1catdead). Consistent with results obtained for NTHL1, both variants of Ntg1 impair cell growth, but only the wild-type protein causes accumulation of double-strand breaks and chromosome loss. We screened a panel of DNA repair mutants for resistance/sensitivity to overexpression of wild-type Ntg1 or ntg1catdead. This analysis identified several cellular pathways that protect cells from Ntg1-induced damage, providing insight into interplay between DNA repair pathways. Finally, we identified a link to sumoylation and probed how this post-translational modification could contribute to regulation of Ntg1 function. This study describes a budding yeast system to understand how cells regulate and respond to dysregulation of the BER pathway. Take AwayO_LIOverexpression of a base excision DNA repair protein impairs cell growth C_LIO_LIOverexpression of a base excision DNA repair protein can cause DNA damage C_LIO_LIMultiple mechanisms cause DNA damage from overexpression of a repair protein C_LIO_LIDNA repair pathways functionally interact to protect cells from DNA damage C_LIO_LIPrecise regulation of the activity of DNA repair proteins is critical C_LI