The Auto-Regulation of ATL2 E3 Ubiquitin Ligase Plays an Important Role in the Immune Response against Alternaria brassicicola in Arabidopsis thaliana

Abstract

The ubiquitin/26S proteasome system is a crucial regulatory mechanism that governs various cellular processes in plants, including signal transduction, transcriptional regulation, and responses to biotic and abiotic stressors. Our study shows that the RING-H2-type E3 ubiquitin ligase, Arabidopsis Tóxicos en Levadura 2 (ATL2), is involved in response to fungal pathogen infection. Under normal growth conditions, the expression of the ATL2 gene is low, but it is rapidly and significantly induced by exogenous chitin. Additionally, ATL2 protein stability is markedly increased via chitin treatment, and its degradation is prolonged when 26S proteasomal function is inhibited. We found that an atl2 null mutant exhibited higher susceptibility to Alternaria brassicicola, while plants overexpressing ATL2 displayed increased resistance. We also observed that the hyphae of A. brassicicola were strongly stained with trypan blue staining, and the expression of A. brassicicola Cutinase A (AbCutA) was dramatically increased in atl2. In contrast, the hyphae were weakly stained, and AbCutA expression was significantly reduced in ATL2-overexpressing plants. Using bioinformatics, live-cell confocal imaging, and cell fractionation analysis, we revealed that ATL2 is localized to the plasma membrane. Further, it is demonstrated that the ATL2 protein possesses E3 ubiquitin ligase activity and found that cysteine 138 residue is critical for its function. Moreover, ATL2 is necessary to successfully defend against the A. brassicicola fungal pathogen. Altogether, our data suggest that ATL2 is a plasma membrane-integrated protein with RING-H2-type E3 ubiquitin ligase activity and is essential for the defense response against fungal pathogens in Arabidopsis.