4.2 m6A methylation regulated the expression of genes
m6A primarily occurs in highly expressed genes during BPH infestation (Figure 2a,b). In contrast, m6A mainly occurs in rice genes with low expression levels during viral infection (Zhang et al., 2021a). Responsive m6A modification is sensitive and complex during biological stress, and different stressors may induce m6A methylation in genes with varying expression levels. Through KEGG analyses, we identified differentially expressed transcripts in many pathways containing m6A modifications (Supporting Information: Figure S11). Four component genes in the principal constituents of rice m6A modification machinery can undergo m6A modification. Their relative expression levels showed considerable variations (Figure 3a–d). Thus, m6A machinery can govern the expression of key component genes via m6A methylation, resulting in significant alterations in overall cellular m6A levels in rice when confronted with BPH infestation. This suggest a potential role for m6A in modulating plant defense against insects.
We found differential regulatory patterns of m6A modifications in gene expression across different plant specific BPH response pathways. The m6A modification direction of genes related to the four methylation components, and major phytohormone pathways (which including JA, SA, auxin, GA), Bph , and cellulose synthesis, showed a positive correlation with the target transcript expression (Figure 3-6; Supporting Information: Table S9, S11, S15, S20). No apparent regulatory patterns were found for the hemicellulose synthesis-related genes (Figure 4C; Supporting Information: Table S14). Thus, the levels of m6A modification display specific enrichment patterns depends on the type of metabolic pathways. Based on studies on regulation of m6A modifications in rice by various viruses and fungi (Shi et al., 2019; Zhang et al., 2021a), our findings suggest a potential role for m6A in modulating plant defense against insects.
4.3 m6A methylation regulated plant defense and growth upon insect infestation
Growth reduction is commonly associated with herbivore attack and defense activation, which is known as the growth-defense trade-off (He et al., 2022). Therefore, plants decide when to grow or defend themselves to optimize their fitness in a changing environment. Research into the molecular mechanisms underlying these growth-defense trade-offs has shown that phytohormones signaling and crosstalk between phytohormones play central roles (Huot et al., 2014; He et al., 2022). Auxin, GA and JA are important growth-and defense-related phytohormones, respectively (Yang et al., 2012). As one of the earliest phytohormones identified, auxin plays an essential role in most aspects of plant growth and development, and also play a vital role in plant defense against diverse pathogens (Spaepen & Vanderleyden, 2011). InArabidopsis , the core JA-responsive basic helix-loop-helix transcription factor MYC2-mediated repression expression of the growth regulator PLT integrates JA action into the auxin pathway in regulating root meristem activity and stem cell niche maintenance (Chen et al., 2011). BPH attack activates GA catabolism in rice plants via two GA catabolism genes, that is, GA2ox3 and GA2ox7 , contribute to BPH-induced growth restriction and are upregulated by JA signaling. The JA-responsive transcription factor MYC2 binds to the promoters of GA2ox3 and GA2ox7 to regulate their expression. Therefore, the MYC2-GA2ox module regulates the growth-defense trade-off when BPH attacks rice, which provides a mechanism for phytohormone crosstalk (Jin et al., 2023).
Although the overall m6A methylation of rice was constrained by BPH damage (Figure 1f,g), JA signaling pathway-related genes were altered and accompanied by a predominance of upregulated transcripts and up-directed m6A methylation sites (Figure 5, Supporting Information: Table S15). Some key genes responsible for rice growth pathways have shown distinct m6A-modification directions from JA pathway. This was reflected in attenuated m6A-modification levels of auxin and GA biosynthesis-related genes under BPH attack, which contributed to the downregulated expression of these transcripts. The rice GA catabolism process was activated after BPH infection, and the significantly upregulated OsGA2ox3 , OsWRKY71 , andOsCRY1b genes displayed a positive correlation between m6A modifications and gene expression (Figure 6, Supporting Information: Table S20-S22). This indicated that plant defense was activated but growth was suppressed during BPH attack, this growth–defense trade-off was likely regulated by m6A methylation-mediated Auxin, GA, and JA pathways in BPH-infested rice.
Overall, m6A modification levels were repressed under BPH infestation, with m6A primarily modifying highly expressed genes. Here, we proposed a simplified model for rice m6A RNA methylation modulation in key defense and growth pathways induced by BPH infestation (Figure 7). m6A modification was positively correlated with the expression of key genes, mainly in insect–stress response and growth pathways in rice, and may have contributed to plants prioritizing defense overgrowth during BPH attack.