Identification of plants’ functional counterpart of the metazoan mediator of DNA Damage checkpoint 1

Induction of DNA damage triggers rapid phosphorylation of the histone H2A.X (γH2A.X). In animals, mediator of DNA damage checkpoint 1 (MDC1) binds γH2A.X through a tandem BRCA1 carboxyl-terminal (tBRCT) domain and mediates recruitment of downstream effectors of DNA damage response (DDR). However, readers of this modification in plants have remained elusive. We show that from the Arabidopsis BRCT domain proteome, BCP1-4 proteins with tBRCT domains are involved in DDR. Through its tBRCT domain BCP4 binds γH2A.X in vitro and localizes to DNA damage-induced foci in an H2A.X-dependent manner. BCP4 also contains a domain that interacts directly with NBS1 and thus acts as a functional counterpart of MDC1. We also show that BCP1, that contains two tBRCT domains, co-localizes with γH2A.X but it does not bind γH2A.X suggesting functional similarity with human PAXIP1. A phylogenetic analysis supports that PAXIP1 and MDC1 in metazoa and their plant counterparts evolved independently from common ancestors with tBRCT domains. Collectively, our study reveals missing components and provides mechanistic and evolutionary insights into plant DDR. Synopsis This study identifies BRACT domain protein 4 (BCP4) as the functional equivalent of Mediator of DNA Damage Checkpoint 1 in plants. A phylogenetic study establishes the criteria defining the identity of this family of proteins in eukaryotes. The tandem-BRCT domain of BCP4 binds phosphorylated H2A.X (γH2A.X) histone variant in vitro. In response to DNA damage, BCP4 co-localizes with γH2A.X foci in vivo. Plant BCP4 has evolved independently from the metazoan MDC1 and has acquired distinct sequence motifs. BCP4 directly binds to NBS1 of the MRN complex through a plant-specific sequence motif. This study identifies BRACT domain protein 4 (BCP4) as the functional equivalent of Mediator of DNA Damage Checkpoint 1 in plants. A phylogenetic study establishes the criteria defining the identity of this family of proteins in eukaryotes.