Background Cucumber green mottle mosaic virus (CGMMV) causes substantial global losses in cucurbit crops, especially watermelon. N6-methyladenosine (m6A) methylation in RNA is one of the most important post-transcriptional modification mechanisms in eukaryotes. It has been shown to have important regulatory functions in some model plants, but there has been no research regarding m6A modifications in watermelon. Results We measured the global m6A level in resistant watermelon after CGMMV infection using a colorimetric method. And the results found that the global m6A level significantly decreased in resistant watermelon after CGMMV infection. Specifically, m6A libraries were constructed for the resistant watermelon leaves collected 48 h after CGMMV infection and the whole-genome m6A-seq were carried out. Numerous m6A modified peaks were identified from CGMMV-infected and control (uninfected) samples. The modification distributions and motifs of these m6A peaks were highly conserved in watermelon transcripts but the modification was more abundant than in other reported crop plants. In early response to CGMMV infection, 422 differentially methylated genes (DMGs) were identified, most of which were hypomethylated, and probably associated with the increased expression of watermelon m6A demethylase gene ClALKBH4B. Gene Ontology (GO) analysis indicated quite a few DMGs were involved in RNA biology and stress responsive pathways. Combined with RNA-seq analysis, there was generally a negative correlation between m6A RNA methylation and transcript level in the watermelon transcriptome. Both the m6A methylation and transcript levels of 59 modified genes significantly changed in response to CGMMV infection and some were involved in plant immunity. Conclusions Our study represents the first comprehensive characterization of m6A patterns in the watermelon transcriptome and helps to clarify the roles and regulatory mechanisms of m6A modification in watermelon in early responses to CGMMV.
Plant carotenoid cleavage oxygenase (CCO) is an enzyme that catalyzes the synthesis of carotenoids and participates in many important physiological functions. The plant CCOs exist in two forms, namely carotenoid cleavage dioxygenase (CCD) and nine-cis epoxide carotenoid dioxygenase (NCED). Although studies have shown that this gene family has been identified in many species, such as Arabidopsis, grape, and tomato, the evolutionary origin of the CCO family and the expression pattern of pepper genes in response to H2O2 and other abiotic stresses are still unclear. In this study, we used the bioinformatics method to identify and analyze the members of the CCO gene family from pepper and other 13 plants from lower to higher plant species based on the whole genome sequence. A total of 158 CCO genes were identified in different plant species and further divided into two groups (e.g., groups I and II). The former was subdivided into CCD7 and CCD8 and have independent evolutionary origins, respectively, while the latter was subdivided into CCD1, CCD4, CCD-like, and NCED, which may have come from a common ancestor. In addition, the results of RNA-seq showed that the expression patterns of pepper CaCCO genes were different in the tissues tested, and only few genes were expressed at high levels such as CaCCD1a, CaCCD4a, CaNCED3, and CaCCD1b. For hydrogen peroxide (H2O2) and other abiotic stresses, such as plant hormones, heat, cold, drought, and NaCl treatments, induction of about half of the CaCCO genes was observed. Moreover, the expression patterns of CaCCOs were further investigated under heat, cold, drought, and NaCl treatments using quantitative real-time PCR (qRT-PCR), and most members were responsive to these stresses, especially some CaCCOs with significant expression changes were identified, such as CaCCD4c, CaCCD-like1, CaCCD8, and CaCCD1b, suggesting the important roles of CaCCOs in abiotic stress responses. All these results will provide a valuable analytical basis for understanding the evolution and functions of the CCO family in plants.
Heat-shock protein 20s (HSP20) were initially shown to play a role during heat shock stress; however, recent data indicated that HSP20 proteins are also involved in abiotic stress in plants. Watermelon is known to be vulnerable to various stressors; however, HSP20 proteins have yet to be investigated and characterized in the watermelon. In a previous study, we identified a negative regulator of salt stress response from watermelon: ClHSP22.8, a member of the HSP20 family. Quantitative real-time PCR (qRT-PCR) and promoter::β-glucuronidase (GUS) analysis revealed that ClHSP22.8 was expressed widely in a range of different tissues from the watermelon, but particularly in the roots of 7-day-old seedlings and flowers. Furthermore, qRT-PCR and GUS staining showed that the expression of ClHSP22.8 was significantly repressed by exogenous abscisic acid (ABA) and salt stress. The over-expression of ClHSP22.8 in Arabidopsis lines resulted in hypersensitivity to ABA and reduced tolerance to salt stress. Furthermore, the expression patterns of key regulators associated with ABA-dependent and independent pathways, and other stress-responsive signaling pathways, were also repressed in transgenic lines that over-expressed ClHSP22.8. These results indicated that ClHSP22.8 is a negative regulator in plant response to salt stress and occurs via ABA-dependent and independent, and other stress-responsive signaling pathways.
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