To maintain genome integrity, cells have developed protection mechanisms, called checkpoints, in response to DNA damage insults. In Saccharomyces cerevisiae, Rad53 protein kinase is one of the major actors in these mechanisms, and its activation triggers several cellular responses such as cell cycle delay, replication delay, transcription modifications, activation of DNA repair pathways… Using an hyperactivative allele of RAD53, we identified HUG1, as one of the most induced gene in a transcriptomic analysis upon RAD53 pathway activation. However Hug1’s functions remains elusive.To better understand Hug1’s functions in DNA damage response, we searched for physical partners and identified Rnr2 and Rnr4 proteins, which are the two small subunits of Ribonucleotide Reductase (RNR). The RNR is an enzymatic complex that catalyses nucleotide reduction, a step limiting for dNTPs synthesis. We next experimentally tackled the Hug1-RNR interaction using various methods. We showed so that Hug1 is a small intrinsically disordered protein able to interact physically with the small RNR subunit and that at least eleven amino acids in Hug1 are involved in this interaction. During our investigations, we observed that C-terminal tagging of Rnr2 sensitizes strains to genotoxics stress and that this sensitivity was suppressed when HUG1’s function is abrogated. Hence, we showed that Hug1 is a negative RNR regulator and propose a model for Hug1’s function.
