LAUSR

Significance Topoisomerases are crucial for genome maintenance and are targets for several chemotherapeutic agents. While anticancer drugs targeting topoisomerases can lead to secondary malignancies, there have been no descriptions of genetic defects in topoisomerases having roles in cancer development. Here we show that a somatic topoisomerase IIα mutation found in human tumors results in a mutator phenotype. We show that this mutation and the concomitant mutational signature, which we call ID_TOP2α, are associated with genomic rearrangements and with potentially oncogenic indel mutations in known driver genes. Our results shed new light on topoisomerase IIα function, on repair of trapped cleavage complexes, and on a likely oncogenic role for topoisomerases. Topoisomerases nick and reseal DNA to relieve torsional stress associated with transcription and replication and to resolve structures such as knots and catenanes. Stabilization of the yeast Top2 cleavage intermediates is mutagenic in yeast, but whether this extends to higher eukaryotes is less clear. Chemotherapeutic topoisomerase poisons also elevate cleavage, resulting in mutagenesis. Here, we describe p.K743N mutations in human topoisomerase hTOP2α and link them to a previously undescribed mutator phenotype in cancer. Overexpression of the orthologous mutant protein in yeast generated a characteristic pattern of 2- to 4-base pair (bp) duplications resembling those in tumors with p.K743N. Using mutant strains and biochemical analysis, we determined the genetic requirements of this mutagenic process and showed that it results from trapping of the mutant yeast yTop2 cleavage complex. In addition to 2- to 4-bp duplications, hTOP2α p.K743N is also associated with deletions that are absent in yeast. We call the combined pattern of duplications and deletions ID_TOP2α. All seven tumors carrying the hTOP2α p.K743N mutation showed ID_TOP2α, while it was absent from all other tumors examined (n = 12,269). Each tumor with the ID_TOP2α signature had indels in several known cancer genes, which included frameshift mutations in tumor suppressors PTEN and TP53 and an activating insertion in BRAF. Sequence motifs found at ID_TOP2α mutations were present at 80% of indels in cancer-driver genes, suggesting that ID_TOP2α mutagenesis may contribute to tumorigenesis. The results reported here shed further light on the role of topoisomerase II in genome instability.