Transcriptome analysis reveals defense-related genes and pathways against Xanthomonas campestris pv. vesicatoria in pepper (Capsicum annuum L.)

Abstract

Bacterial spot (BS), incited by Xanthomonas campestris pv. vesicatoria (Xcv), is one of the most serious diseases of pepper. For a comparative analysis of defense responses to Xcv infection, we performed a transcriptomic analysis of a susceptible cultivar, ECW, and a resistant cultivar, VI037601, using the HiSeqTM 2500 sequencing platform. Approximately 120.23 G clean bases were generated from 18 libraries. From the libraries generated, a total of 38,269 expressed genes containing 11,714 novel genes and 11,232 differentially expressed genes (DEGs) were identified. Functional enrichment analysis revealed that the most noticeable pathways were plant-pathogen interaction, MAPK signaling pathway—plant, plant hormone signal transduction and secondary metabolisms. 1,599 potentially defense-related genes linked to pattern recognition receptors (PRRs), mitogen-activated protein kinase (MAPK), calcium signaling, and transcription factors may regulate pepper resistance to Xcv. Moreover, after Xcv inoculation, 364 DEGs differentially expressed only in VI037601 and 852 genes in both ECW and VI037601. Many of those genes were classified as NBS-LRR genes, oxidoreductase gene, WRKY and NAC transcription factors, and they were mainly involved in metabolic process, response to stimulus and biological regulation pathways. Quantitative RT-PCR of sixteen selected DEGs further validated the RNA-seq differential gene expression analysis. Our results will provide a valuable resource for understanding the molecular mechanisms of pepper resistance to Xcv infection and improving pepper resistance cultivars against Xcv.

Identification of Key Transcription Factors Related to Bacterial Spot Resistance in Pepper through Regulatory Network Analyses

“…In contrast, it turns out that the 4 TF candidates, Capana00g004634 (ERF family), Capana09g000015 (C3H family), Capana11g000364 (TALE family), and Capana03g003818 (HSF family) in our study are different from the aforementioned differentially expressed TFs in the original paper [32] even though they were all considered as candidate genes related to Xcv resistance in pepper in the two different studies. In our opinion, different conclusions regarding candidates of resistance genes between our study and the original paper [32] could reflect variations in outcomes caused by the application of different prediction methodologies. In the original article [32], the candidates of resistance genes were predicted based on DEGs that occurred in ECW and VI037601 at the time points (6hpi and 24 hpi) post Xcv infection, whereas candidates of resistance genes were predicted in our study through WGCNA in combination with GENIE3 network constructed using almost all genes instead of DEGs.…”

“…In our opinion, different conclusions regarding candidates of resistance genes between our study and the original paper [32] could reflect variations in outcomes caused by the application of different prediction methodologies. In the original article [32], the candidates of resistance genes were predicted based on DEGs that occurred in ECW and VI037601 at the time points (6hpi and 24 hpi) post Xcv infection, whereas candidates of resistance genes were predicted in our study through WGCNA in combination with GENIE3 network constructed using almost all genes instead of DEGs. Both results could be complementary with each other, thereby providing more candidates for pepper breeding even though the prediction accuracy still awaits future validation.…”

“…Public RNA-seq data sets are available from NCBI’s SRA (Sequence Read Archive) database with accession number PRJNA693027 [ 32 ]. The detailed procedures for generating RNA-seq data from two bell pepper cultivars, VI037601 and ECW, representing resistance and susceptibility to Xcv infection, respectively, were previously described [ 32 ].…”

“…Public RNA-seq data sets are available from NCBI’s SRA (Sequence Read Archive) database with accession number PRJNA693027 [ 32 ]. The detailed procedures for generating RNA-seq data from two bell pepper cultivars, VI037601 and ECW, representing resistance and susceptibility to Xcv infection, respectively, were previously described [ 32 ]. Briefly, two cultivars were grown at 25 °C/20 °C day/dark time and 16 h/8 h light/dark cycle with an approximately 60% relative humidity in standard glasshouse conditions.…”

“…The differentially expressed genes (DEGs) in VI037601 and early Calwonder (ECW), representing resistance and susceptibility to Xcv infection, respectively, at 0 h, 6 h, and 24 h post Xcv infection, have already been characterized by Gao et al [ 32 ]. However, it is still unclear how genes are differentially expressed between VI037601 and ECW at 0 h, 6 h, and 24 h post Xcv infection.…”