The nucleolar protein GNL3 prevents resection of stalled replication forks

Faithful DNA replication requires specific proteins that protect replication forks and so prevent the formation of DNA lesions that may damage the genome. Identification of new proteins involved in this process is essential to understand how DNA lesions accumulate in cancer cells and how they tolerate them. Here, we show that human GNL3/nucleostemin, a GTP-binding protein localized mostly in the nucleolus and highly expressed in cancer cells, prevents nuclease-dependent resection of nascent DNA in response to replication stress. We demonstrate that inhibiting origin firing reduces resection. This suggests that the heightened replication origin activation observed upon GNL3 depletion largely drives the observed DNA resection probably due to the exhaustion of the available RPA pool. We show that GNL3 and DNA replication initiation factor ORC2 interact in the nucleolus and that the concentration of GNL3 in the nucleolus is required to limit DNA resection. We propose that the control of origin firing by GNL3 through the sequestration of ORC2 in the nucleolus is critical to prevent nascent DNA resection in response to replication stress. imageGNL3 limits the number of fired replication origins by sequestering ORC2 within the nucleolus. In the absence of GNL3, increased replication origin firing triggers nuclease-mediated resection of newly synthesized DNA in response to replication stress.GNL3 is a GTP-binding protein primarily found within the nucleolus.The nucleolar accumulation of GNL3 limits the number of replication origins that fire, likely through the sequestration of ORC2.When GNL3 is deficient or fails to accumulate in the nucleolus, it triggers nuclease-mediated resection of newly synthesized DNA in response to replication stress. GNL3 limits the number of fired replication origins by sequestering ORC2 within the nucleolus. In the absence of GNL3, increased replication origins firing triggers nuclease-mediated resection of newly synthesized DNA in response to replication stress.image

Automated summary

This study identifies a protein called GNL3 that protects DNA integrity during replication by limiting DNA resection. It interacts with the DNA replication initiation factor ORC2 in the nucleolus to control the number of activated replication origins and prevent DNA resection.