Transcriptome-wide N6-methyladenosine (m6A) methylation in watermelon under CGMMV infection

Abstract

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.

AlkB RNA demethylase homologues and N6‐methyladenosine are involved in Potyvirus infection

“…It has been suggested to be part of a novel antiviral system because genetic depletion of a plant m 6 A demethylase negatively affects alfalfa mosaic virus accumulation and movement (Martínez‐Pérez et al, 2017, 2021). The overall plant m 6 A level has been reported to be altered by RNA viruses of the genera Tobamovirus , Bymovirus, Tenuivirus , and Fijivirus (He et al, 2021; Li et al, 2018; Zhang, Zhuang, et al, 2021; Zhang, Wang, et al, 2021). Despite these recent studies, the biological significance of internal RNA methylation and m 6 A in plant virus infection and antiviral immunity is poorly understood.…”

m6A‐mediated regulation of crop development and stress responses

“…In watermelon, the infection by cucumber green mottle mosaic virus (CGMMV) leads to the decrease in global m 6 A level and the increase in expression of m 6 A demethylase gene ClALKBH4B (He et al, 2021). Some m 6 A-modified transcripts related to plant immunity display altered m 6 A enrichments and transcript levels in response to CGMMV infection.…”

“…In addition, when the m 6 A methyltransferase genes LsMETTL3 and LsMETTL14 from small brown planthopper (SBPH), the insect vector of RBSDV, were silenced, the titre of RBSDV in the midgut cells of SBPHs increased significantly, suggesting a negative correlation between the overall m 6 A levels and the virus replication (Tian et al ., 2021 ). In watermelon, the infection by cucumber green mottle mosaic virus (CGMMV) leads to the decrease in global m 6 A level and the increase in expression of m 6 A demethylase gene ClALKBH4B (He et al ., 2021 ). Some m 6 A‐modified transcripts related to plant immunity display altered m 6 A enrichments and transcript levels in response to CGMMV infection.…”

“…Some m 6 A‐modified transcripts related to plant immunity display altered m 6 A enrichments and transcript levels in response to CGMMV infection. It is probably that ClALKBH4B modulates the expression of the defence genes via m 6 A methylation, thereby participating in the regulation of watermelon against CGMMV infection (He et al ., 2021 ).…”

Insights into the Molecular Mechanisms of the N6-Methyladenosine (m6A) Methylation Machinery in the Regulation of the Infection Cycle of RNA Plant Viruses

“…In this sense, it has been shown that the m 6 A-level is reduced after TMV infection in Nicotiana tabacum plants . In watermelon, global m 6 A levels are reduced in response to cucumber green mottle mosaic virus (CGMMV) infection implying that a global hypomethylation of m 6 A likely activates watermelon defense responses early in CGMMV infection (He et al, 2021). Furthermore, infected plants showed…”