Crown gall disease, caused by the pathogenic bacterium Agrobacterium tumefaciens, is responsible for extensive economic losses in orchards. Cherry rootstock ‘CDR-1’ (Prunus mahaleb) shows high resistance but the mechanism remains unclear. Here, we examined the morphology of pathogen-infected root neck surface, determined the activity of 10 defense-related enzymes and the content of salicylic acid (SA) and jasmonic acid (JA), and also applied transcriptome analysis, transient expression and transgenic verification to explore the crown gall resistance genes in ‘CDR-1’ plants. In our study, peroxidase increased in the first 10?days, while phenylalanine ammonialyase and lipoxygenase increased in the first 15?days post-infection. Four key enzymes in the AsA-GSH cycle also responded, to a certain extent; although JA content increased significantly after the treatment, the SA content did not. In a follow-up transcriptome analysis, the differentially expressed genes Pm4CL2, PmCYP450, PmHCT1, PmHCT2, and PmCAD were up-regulated. Based on the above results, we focused on the lignin biosynthetic pathway, and further measured lignin content, and found it increased significantly. The Pm4CL2 gene was used to conduct transient expression and transgenic experiments to verify its function in crown gall disease resistance. It showed the relative expression of the treatment group was almost 14-fold that of the control group at 12?h post-treatment. After the infection treatment, clear signs of resistance were found in the transgenic lines; this indicated that under the higher expression level and earlier activation of Pm4CL2, plant resistance was enhanced. The crown gall resistance of ‘CDR-1’ is likely related to the lignin biosynthetic pathway, in which Pm4CL2 functions crucially during the plant defense response to the pathogen A. tumefaciens. The results thus offer novel insights into the defense responses and resistance mechanism of cherry rootstock ‘CDR-1’ against crown gall disease.
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