JA signal-mediated immunity of Dendrobium catenatum to necrotrophic Southern Blight pathogen

Abstract

Background: Dendrobium catenatum belongs to the Orchidaceae, and is a precious Chinese herbal medicine. In the past 20 years, D. catenatum industry has developed from an endangered medicinal plant to multi-billion dollar grade industry. The necrotrophic pathogen Sclerotium delphinii has a devastating effection on over 500 plant species, especially resulting in widespread infection and severe yield loss in the process of large-scale cultivation of D. catenatum. It has been widely reported that Jasmonate (JA) is involved in plant immunity to pathogens, but the mechanisms of JA-induced plant resistance to S. delphinii are unclear.

Results: In the present study, the role of JA in enhancing D. catenatum resistance to S. delphinii was investigated. We identified 2 COI1, 13 JAZ, and 12 MYC proteins in D. catenatum genome. Subsequently, systematic analyses containing phylogenetic relationship, gene structure, protein domain, and motif architecture of core JA pathway proteins were conducted in D. catenatum and the newly characterized homologs from its closely related orchid species Phalaenopsis equestris and Apostasia shenzhenica, along with the well-investigated homologs from Arabidopsis thaliana and Oryza sativa. Public RNA-seq data were investigated to analyze the expression patterns of D. catenatum core JA pathway genes in various tissues and organs. Transcriptome analysis of MeJA and S. delphinii treatment showed exogenous MeJA changed most of the expression of the above genes, and several key members, including DcJAZ1/2/5 and DcMYC2b, are involved in enhancing defense ability to S. delphinii in D. catenatum.

Conclusions: The findings indicate exogenous MeJA treatment affects the expression level of DcJAZ1/2/5 and DcMYC2b, thereby enhancing D. catenatum resistance to S. delphinii. This research would be helpful for future functional identification of core JA pathway genes involved in breeding for disease resistance in D. catenatum.

Keywords: COI1; Dendrobium catenatum; JAZ; Jasmonate; MYC transcription factor; Sclerotium delphinii.

Fig. 1

Effect of exogenous MeJA pre-treatment on D. catenatum resistance to S. delphinii. A Disease symptoms of D. catenatum plantlets pre-treated with 1 mM MeJA following inoculation with S. delphinii at 60 and 240 hpi. Bar = 1 cm. B Disease indexes of the MeJA pre-treated D. catenatum were determined from 24 hpi (1 dpi) to 240 hpi (10 dpi). The values are the means ± SD; n = 10

Fig. 2

Phylogenetic relationships, architecture of conserved protein domains and motifs, and gene structure in COI genes of D. catenatum, P. equestris, A. shenzhenica, A. thaliana, and O. sativa. A The phylogenetic tree of COI proteins. It was constructed according to the NJ method by MEGA 7.0 with 1000 bootstrap replicates. B The conserved domain and motif composition of COI proteins. The Solid triangle with orange colors represented the F-box domain. The Solid boxes with different colors represented different motif, and the legend was given at the right of figure. C Exon–intron structure of COI genes. Green boxes represents exons and black lines represent introns

Fig. 3

Phylogenetic analysis of 89 TIFY proteins from D. catenatum, P. equestris, A. shenzhenica, A. thaliana, and O. sativa. The phylogenetic tree was constructed according to the NJ method by MEGA 7.0 with 1000 bootstrap replicates. DcTIFYs, PeTIFYs, AsTIFYs, AtTIFYs, and OsTIFYs were labeled with red star, yellow square, green hook, blue triangle, pink circle, respectively. The eight groups with different colors represent eight clades

Fig. 4

Phylogenetic relationships, architecture of conserved protein domains and motifs, and gene structure in JAZ genes of D. catenatum, P. equestris, A. shenzhenica, A. thaliana, and O. sativa. A The phylogenetic tree of JAZ proteins. Different color boxes represent five different clades. B The conserved domain and motif composition of JAZ proteins. The Solid triangle with different colors represented different conserved protein domains (TIFY, Jas, CMID, and EAR). The Solid boxes with different colors represented different motifs, and the legend was given at the right of the figure. C Exon–intron structure of JAZ genes. Green boxes represent exons and black lines represents introns. Sequence logos of the TIFY (D) and Jas (E) domains from D. catenatum JAZ proteins

Fig. 5

Phylogenetic analysis of 57 MYC proteins from D. catenatum, P. equestris, A. shenzhenica, A. thaliana, and O. sativa. The phylogenetic tree was constructed according to the NJ method by MEGA 7.0 with 1000 bootstrap replicates. DcTIFYs, PeTIFYs, AsTIFYs, AtTIFYs, and OsTIFYs were labeled with red star, yellow square, green hook, blue triangle, pink circle, respectively. The five groups with different colors represent five clades

Fig. 6

Phylogenetic relationships, architecture of conserved protein domains and motifs, and gene structure in MYC genes of D. catenatum, P. equestris, A. shenzhenica, A. thaliana, and O. sativa. A The phylogenetic tree of MYC proteins. Different color boxes represent five different clades. B The conserved domain and motif composition of MYC proteins. The Solid triangle with orange and light blue represented bHLH domain and bHLH-MYC_N domain, respectively. The Solid boxes with different colors represented different motif and the legend was given at the right of figure. C Exon–intron structure of MYC genes. Green boxes represent exons and black lines represent introns. D Sequence alignment of JID, TAD, basic, and HLH domain of DcMYC2a, DcMYC2b, DcMYC2c, DcMYC2d, AtMYC2, AtMYC3, AtMYC4, and AtMYC5 proteins. Sequence alignment was performed using ClustalX. Red triangles represented amino acid residues involved in the AtMYC3-JAZ interaction. Blue triangles represented residues that are related to specific DNA recognition in MYC2. Orange triangles and black triangles represented residues that are required for MYC2 dimer formation and tetramer formation, respectively

Fig. 7

Expression profiles of JA signaling pathway genes in different tissues and organs in D. catenatum. A Expression patterns of DcCOI genes in ten D. catenatum tissues and organs. B Expression patterns of DcJAZ genes in ten D. catenatum tissues and organs. C Expression patterns of DcMYC genes in ten D. catenatum tissues and organs. Lf: leaves, Ro: roots, Gr: green root tip, Wr: the white part of roots, St: stems, Fb: flower buds, Se: sepals, Lb: labellum (lip), Po: pollinia, and Gs, gynostemium (column). The log₂ transformations of the expression values were used to generate the heat map with TBtools software

Fig. 8

Expression profiles of JA signaling pathway genes response to S. delphinii after MeJA pre-treated. Heat map showing expression pattern of DcCOI, DcJAZ, DcMYC, and JA pathway marker genes (DcPR3, DcLOX2, DcTAT3, and DcVSP2) in leaves under different treatments. CK: control. P1: 24 h post inoculated with S. delphinii. JA: pre-treated by MeJA for 4 h and then inoculated with sterile distilled water for 24 h. P1 + JA: pre-treated by MeJA for 4 h and then inoculated with S. delphinii for 24 h. The log₂ transformations of the expression values were used to generate the heat map with TBtools software

Fig. 9

Expression of 11 JA signaling pathway genes response to S. delphinii after MeJA-pretreated through RT-qPCR assay. CK: control. P1: 24 h post inoculated with S. delphinii. JA: pre-treated by MeJA for 4 h and then inoculated with sterile distilled water for 24 h. P1 + JA: pre-treated by MeJA for 4 h and then inoculated with S. delphinii for 24 h.The actin gene of D. catenatum was used as an internal control. The error bars indicate SD from three independent experiments. The * and ** show the significant difference at P < 0.05 and P < 0.01 compared with the CK by Student’s test, respectively