The Arabidopsis deNADding enzyme DXO1 modulates the plant immunity response.
The DXO family proteins, highly conserved among eukaryotes, are involved in mRNA 5'-end quality control (5'QC) and also remove non-canonical NAD+ cap from RNA (deNADding). DXO1 protein, the only DXO homolog in Arabidopsis, due to its plant-specific features has a strong deNADding enzymatic activity but no apparent role in 5' QC. Despite its significant contribution to plant RNA metabolism, a direct role for DXO1 enzymatic activity in the cell has not yet been demonstrated. Notably, all molecular and morphological changes observed so far in dxo1 mutant plants depended on the plant-specific N-terminal domain of the protein. Our investigation into the role of DXO1 in response to biotic stress, specifically its susceptibility to Pseudomonas syringae pv. tomato (Pst) DC3000 infection, unexpectedly revealed the importance of DXO1 enzymatic activity in the plant immune response. We observed that the dxo1-2 knockout mutant and transgenic dxo1-2 lines expressing a DXO1 variant either catalytically inactive (DXO1(E394A/D396A)) or lacking the N-terminal domain (DXO1(deltaN194)) exhibited enhanced resistance to Pst, accompanied by marked changes in the expression of key pathogenesis markers. Also, other markers of plant immunity, such as callose deposition and production of reactive oxygen species, were strongly induced by two pathogen effectors, elf18 and flg22, in the tested mutant lines. These results strongly suggest that both DXO1 features, the N-terminal domain and its catalytic center, contribute to the regulation of plant immunity. In conclusion, this is the first observation revealing the involvement of the DXO1 enzymatic activity in plant physiology. Founded by National Science Centre Grant, Poland (UMO-2014/13/B/NZ3/00405, UMO-2018/29/B/NZ3/01980, UMO-2021/40/Q/NZ1/00014). Overall design: Four replicates of Col-0 and dxo1-2 mutant plants were treated with Pseudomonas syringae pv. tomato DC3000 or a mock control, and RNA samples were subjected to 3'RNA-seq.