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

Over 50 hereditary degenerative disorders are caused by expansions of short tandem DNA repeats (STRs). (GAA) repeat expansions are responsible for Friedreich's ataxia as well as late-onset cerebellar ataxias (LOCAs). Thus, the mechanisms of (GAA) 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) repeat tract. We found that DNA nicks 5' of the (GAA) 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) repeats, recreating, in a model system, the expansion events that occur in human pedigrees. DNA nicks 3' of the (GAA) 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.