Involvement of AtPolλ in the repair of high salt- and DNA cross-linking agent-induced double strand breaks in arabidopsis

Abstract

DNA polymerase l (Pol l) is the sole member of family X DNA polymerase in plants and plays a crucial role in nuclear DNA damage repair. Here, we report the transcriptional up-regulation of Arabidopsis (Arabidopsis thaliana) AtPoll in response to abiotic and genotoxic stress, including salinity and the DNA cross-linking agent mitomycin C (MMC). The increased sensitivity of atpoll knockout mutants toward high salinity and MMC treatments, with higher levels of accumulation of double strand breaks (DSBs) than wild-type plants and delayed repair of DSBs, has suggested the requirement of Pol l in DSB repair in plants. AtPoll overexpression moderately complemented the deficiency of DSB repair capacity in atpoll mutants. Transcriptional up-regulation of major nonhomologous end joining (NHEJ) pathway genes KU80, X-RAY CROSS COMPLEMENTATION PROTEIN4 (XRCC4), and DNA Ligase4 (Lig4) along with AtPoll in Arabidopsis seedlings, and the increased sensitivity of atpoll-2/atxrcc4 and atpoll-2/atlig4 double mutants toward high salinity and MMC treatments, indicated the involvement of NHEJ-mediated repair of salinity- and MMC-induced DSBs. The suppressed expression of NHEJ genes in atpoll mutants suggested complex transcriptional regulation of NHEJ genes. Pol l interacted directly with XRCC4 and Lig4 via its N-terminal breast cancer-associated C terminus (BRCT) domain in a yeast two-hybrid system, while increased sensitivity of BRCT-deficient Pol l-expressing transgenic atpoll-2 mutants toward genotoxins indicated the importance of the BRCT domain of AtPoll in mediating the interactions for processing DSBs. Our findings provide evidence for the direct involvement of DNA Pol l in the repair of DSBs in a plant genome. © 2013 American Society of Plant Biologists. All Rights Reserved.