Protein UFMylation regulates early events of ribosomal DNA double-stranded break response

The highly repetitive and transcriptionally active ribosomal DNA (rDNA) genes are exceedingly susceptible to genotoxic stress. Induction of DNA double-strand breaks (DSBs) in rDNA repeats is associated with ATM-dependent rDNA silencing and nucleolar reorganization where rDNA is segregated into nucleolar caps. However, the regulatory events underlying this response remain elusive. Here, we identify protein UFMylation as essential for rDNA damage response in human cells. We further show the only UFM1-E3-ligase UFL1 and its binding partner DDRGK1 localize to nucleolar caps upon rDNA damage, and that UFL1 loss impairs ATM activation and rDNA transcriptional silencing, leading to reduced rDNA segregation. A first-ever analysis of nuclear and nucleolar UFMylation targets in response to DSBs induction further identified key DNA repair factors including ATM, in addition to chromatin and actin network regulators. Taken together, our data provides the first evidence of an essential role for UFMylation in orchestrating rDNA DSB repair.