Mechanism of ferulic acid-induced resistance to soft rot disease in kiwifruit based on transcriptomics anal ysis

【Objective】The study aimed to try to find natural and safe plant growth-promoting agents in order to effectively control the post-harvest soft rot disease of kiwifruit (Actinidia chinensis) caused by Botryosphaeria dothidea. Previously, some researchers have stated that ferulic acid (FA), as a natural phenolic acid, can induce the expression levels of related genes in apples and tomatoes, thereby enhancing their disease resistance. Moreover, the application of transcriptomics technology in post- harvest preservation of fruits and vegetables has been widely reported. Therefore, in this study, using transcriptomics technology, the influence of ferulic acid on the related genes of kiwifruit fruits for post-harvest soft rot disease resistance was analyzed with intention to provide a theoretical basis for the preliminary exploration of the molecular mechanism of ferulic acid inducing post-harvest soft rot disease resistance in kiwifruit fruits.【Methods】Hongyang kiwifruit (Actinidia chinensis) was used as the test material. The fruits were immersed in 0.8 g·L-1 FA solution for 10 min, and the same amount of sterile water was used as the control for the same time. Then, B. dothidea was inoculated, and the lesion diameters were measured daily. The tissue samples of the FA treatment group, inoculation group, FA + inoculation group, control group and blank control group at 0 d after inoculation of B. dothidea were selected for transcriptome sequencing, and RT- qPCR technology was used for verification.【Results】Compared with the inoculation group, the diameter of the disease spots on the kiwi fruits in the FA+ inoculation group was significantly reduced, and the induction effect was significantly increased. The peak value was reached on the 5th day after inoculation, at 18.27%. Through transcriptome sequencing and data analysis of 15 samples, an average of 6.62 Gb of clean data was obtained for each sample, with high data quality. The results of reference genome alignment showed that the sequencing data of this study were highly matched to the kiwi reference genome. The correlation analysis of samples indicated good repeatability within the groups. Through the annotation analysis of transcription factors in the transcriptome data, it was found that the families with a relatively large proportion were bHLH (1643), NAC (1218), MYB_related (1154), ERF (965), WRKY (697), etc. These transcription factors are closely related to plant disease resistance. Compared with the control group, the single treatment group with ferulic acid had 622 genes upregulated and 308 genes downregulated; compared with the inoculation group, the FA+inoculation group had 500 genes upregulated and 95 genes downregulated. The results showed that ferulic acid treatment induced changes in the expression of kiwi-related genes. The GO analysis indicated that ferulic acid treatment had a significant impact on the metabolism and catalytic functions of enzymes during the post-harvest storage of kiwi fruits. The KEGG pathway enrichment results showed that after FA treatment of kiwi fruits, differentially expressed genes (DEGs) were enriched in pathways such as plant-pathogen interaction, secondary metabolism, ascorbic acid and aldehyde acid metabolism, glutathione metabolism, phenylpropanoid biosynthesis, flavonoid biosynthesis, and MAPK signaling pathway. Further research found that FA treatment caused transcriptional reprogramming of kiwi fruits. FA may regulate the expression of genes related to the AsA-GSH cycle, thereby coordinating the regulation of glutathione and ascorbic acid contents to maintain ROS homeostasis. Secondly, FA may also coordinate the regulation of lignin and flavonoid contents by regulating the expression levels of key genes in the phenylpropanoid biosynthesis pathway. Moreover, correlation analysis showed that multiple differentially expressed WRKYs and MYBs were significantly correlated with some key genes of the phenylpropanoid biosynthesis pathway (the AcCHS, AcHCT, AcF5H, and AcCOMT), and Actinidia17942 was significantly positively correlated with the AcCHS, AcHCT, and AcF5H; the Actinidia37434 was significantly correlated with the AcCHS, AcHCT, and AcCOMT. This indicated that Actinidia17942 and Actinidia37434 might synergistically regulate the synthesis of flavonoids and lignins. Furthermore, it was found that most DEGs in the MAPK signaling pathway were upregulated. The protein-protein interaction network (PPI) analysis showed that the Actinidia03761 was the core of the network, and the Actinidia03761 was located in the MAPK signaling pathway. Therefore, it is speculated that the Actinidia03761 may be the core of the regulatory network of FA-induced resistance to post-harvest soft rot disease in kiwi. Finally, RT- qPCR results verified the reliability of the RNA- seq data.【Conclusion】In summary, the treatment with ferulic acid could enhance the post-harvest resistance of kiwifruit to soft rot disease. This might be achieved by regulating the AsA-GSH cycle, phenylpropanoid biosynthesis, and the MAPK signaling pathway, thereby strengthening the defense against B. dothidea. Investigating the regulatory effect of ferulic acid on the related genes for disease resistance in kiwifruit would provide a theoretical basis for effectively controlling postharvest soft rot disease in kiwifruit.