Protein kinase ATR inhibits E3 ubiquitin ligase CRL4PRL1 to stabilize ribonucleotide reductase in response to replication stress

The protein kinase ATR is essential for replication stress responses in all eukaryotes. Ribonucleotide reductase (RNR) catalyzes the formation of deoxyribonucleotide (dNTP), the universal building block for DNA replication and repair. However, the relationship between ATR and RNR is not well understood. Here, we show that ATR promotes the protein stability of RNR in Arabidopsis. Through an activation tagging-based genetic screen, we found that overexpression of TSO2, a small subunit of RNR, partially suppresses the hypersensitivity of the atr mutant to replication stress. Biochemically, TSO2 interacts with PRL1, a central subunit of the Cullin4-based E3 ubiquitin ligase CRL4PRL1, which polyubiquitinates TSO2 and promotes its degradation. ATR inhibits CRL4PRL1 to attenuate TSO2 degradation. Our work provides an important insight into the replication stress responses and a post-translational regulatory mechanism for RNR. Given the evolutionary conservation of the proteins involved, the ATR-PRL1-RNR module may act across eukaryotes. [Display omitted] •Overexpression of TSO2, a subunit of ribonucleotide reductase, suppresses atr•PRL1 directly interacts with TSO2 both in vitro and in vivo•PRL1 functions in CRL4PRL1 to polyubiquitinate TSO2 and promote TSO2 degradation•ATR negatively regulates PRL1 to stabilize TSO2 in response to replication stress Bao et al. show that the evolutionarily conserved E3 ubiquitin ligase CRL4PRL1 mediates polyubiquitination and degradation of the small subunit of ribonucleotide reductase, which catalyzes the formation of dNTP required for DNA replication and repair, identifying mechanisms for replication stress response and post-translational regulation of ribonucleotide reductase.