ABSTRACT Base excision repair (BER) is a critical pathway for repairing damaged DNA bases in cells; however, the mechanisms of protein recruitment and interaction in this pathway remain largely unexplored in higher plants. In this study, we used ‘84K’ poplar ( Populus alba × P. glandulosa ) as the experimental system and applied a low concentration of 5‐aminouracil (5‐AU) to induce DNA base lesions. Through transcriptome analysis and weighted gene co‐expression network analysis (WGCNA), we identified two key BER‐responsive genes: the DNA glycosylase family gene PagDMG6341 and the DNA polymerase δ subunit PagPOLD4 . PagDMG6341 was significantly upregulated during the arrest phase of 5‐AU treatment, whereas PagPOLD4 expression peaked during the subsequent release phase. RNA interference (RNAi) lines for each gene resulted in impaired growth and increased susceptibility to 5‐AU in ‘84K’ poplar, supporting their functional roles in DNA repair and development. To further investigate their potential interaction network, we performed yeast two‐hybrid (Y2H) screening, AlphaFold3‐based structural modelling, confirmatory Y2H, bimolecular fluorescence complementation (BiFC) assays, and luciferase complementation imaging (LCI) assays. These experiments demonstrated that a Transducin/WD40‐repeat‐like scaffold protein (PagWD40) interacts independently with both PagDMG6341 and PagPOLD4. The yeast three‐hybrid (Y3H) assay further showed that PagWD40 functions as a molecular scaffold, linking PagDMG6341 and PagPOLD4 to form a functional complex. This study reveals a new mechanism in which PagWD40 functions as a scaffold protein linking a DNA glycosylase with DNA polymerase δ in the plant BER pathway, thereby providing new insights into the organisation of plant DNA damage repair networks.
Ling et al. (Mon,) studied this question.