Recurrent DNA nicks drive massive expansions of (GAA)n repeats

Abstract

Over 50 hereditary degenerative disorders are caused by expansions of short tandem DNA repeats (STRs). (GAA) n repeat expansions are responsible for Friedreich’s ataxia as well as late-onset cerebellar ataxias (LOCAs). Thus, the mechanisms of (GAA) n repeat expansions attract broad scientific attention. To investigate the role of DNA nicks in this process, we utilized a CRISPR-Cas9 nickase system to introduce targeted nicks adjacent to the (GAA) n repeat tract. We found that DNA nicks 5′ of the (GAA) 100 run led to a dramatic increase in both the rate and scale of its expansion in dividing cells. Strikingly, they also promoted large-scale expansions of carrier- and large normal-size (GAA) n repeats, recreating, in a model system, the expansion events that occur in human pedigrees. DNA nicks 3′ of the (GAA) 100 repeat led to a smaller but significant increase in the expansion rate as well. Our genetic analysis implies that in dividing cells, conversion of nicks into double-strand breaks (DSBs) during DNA replication followed by DSB or fork repair leads to repeat expansions. Finally, we showed that 5′ GAA-strand nicks increase expansion frequency in nondividing yeast cells, albeit to a lesser extent than in dividing cells.