Characterization of DPa and DPb transcription regulators in Arabidopsis thaliana : participation in DNA damage responses

LUCIO SIMONELLI; MARÍA LUJAN SHERIDAN; PAULA CASATI

Congreso; Congreso SAIB 2022; 2022

Because of their sessile condition, plants are continuously exposed to solar radiation. One component ofthe solar spectrum is UV-B radiation (280-315 nm). At high intensities, it can damage lipids, proteins andmost important, DNA. As a consequence, exposure to UV-B can inhibit plant growth and elongation ofprimary roots. If damaged DNA cannot be adequately repaired, cells enter the death cell program. TheG1/S transition is an important checkpoint of the eukaryotic cell division cycle, in which the cells decide tocontinue or to stop dividing. One of the most important regulators is the Retinoblastoma (Rb) protein,which can negatively regulate E2F transcriptional factors. In Arabidopsis thaliana , there are six E2Fproteins (E2Fa – E2Ff), which control the expression of G1/S transition genes. E2Fa-c are called “typicalE2Fs” and they need to heterodimerize with two DP interaction proteins (DPa and DPb) to regulatetranscription of target genes. Different studies in our and other labs showed that the effect of UV-Bradiation on E2F mutants (e2fa , b and c ) exert different responses. These results demonstrate that E2F TFsplay crucial and in some cases antagonist roles in several pathways related to cell division, DNA repairand differentiation. However, little is known about the specific roles of the DP interacting proteins in theseresponses. In this work, we have phenotypically characterized DPa and DPb overexpressing (OE) andmutant lines. We also studied the participation of those proteins in the regulation of cell proliferation andplant growth in response to UV-B exposure. DPa and DPb overexpressing lines showed shorter primaryroots than WT seedlings under control conditions. We used confocal microscopy to analyze the primaryroot meristematic zones from these plants. DPaOE and DPbOE have shorter meristematic zones with fewercells than those from WT primary roots. On the contrary, when primary roots from dpa and dpb mutantswere studied, they looked similar to those from WT seedlings, showing meristematic zones with similarsize as those from WT seedlings. Previously, inhibition of primary root elongation after UV-B irradiation inWT Arabidopsis seedlings was correlated to a decrease in the length and in the number of cells of themeristematic zone. After UV-B exposure, DPaOE and DPbOE primary roots were also shorter than thosefrom WT seedlings; however, the effect of the treatment was similar in the transgenic and the WT plants.When the effect of a single UV-B exposure in dpa and dpb mutants was analyzed, the results showed thatdpb mutants showed a similar inhibition of primary root elongation as WT roots, but after the treatment,dpa roots were longer than those from WT seedlings, showing less sensitivity to UV-B. This result wasconfirmed by the analysis of the primary root meristems, showing those from dpa seedlings larger sizeswith more cells. Finally, we analyzed how programmed cell death is activated after UV-B exposure in thedifferent lines. Interestingly, while all plants showed dead cells in the primary root meristems after UV-Bexposure, both DPaOE and dpb mutants had a lower number of dead cells after UV-B treatment than WT,DPbOE and dpa mutants. Despite this, all plants accumulated similar levels of cyclobutane pyrimidinedimers after UV-B, suggesting that none of these proteins directly participate in DNA repair. Together, ourresults suggest that DPa and DPb are both involved in the UV-B response in roots, having differentfunctions.