Rad27/FEN1 prevents accumulation of Okazaki fragments and ribosomal DNA copy number changes

DNA copy number changes are the most frequent genomic alterations in cancer cells. Here, we demonstrate that Rad27/FEN1, a structure-specific nuclease in budding yeast, plays a crucial role in maintaining the stability of the ribosomal DNA (rDNA) repeats. Severe rDNA instability is observed in the rad27∆ mutant, independently of Fob1-mediated DNA replication fork arrest and DNA double-strand break (DSB) formation in the rDNA. The rad27Δ mutant accumulates Okazaki fragments in the rDNA region, without inducing the formation of detectable DSBs. Similar rDNA instability is observed in DNA ligase^Cdc9-deficient cells. Furthermore, Exonuclease 1 and PCNA partially compensate for the loss of Rad27 in rDNA stabilization. These findings highlight the importance of proper Okazaki fragment maturation in the maintenance of rDNA stability.