ZnO nanoparticles enhances cadmium tolerance by modulating N6-methyladenosine (m6A) level of stress-responsive genes NRT1 and GM35E in vegetable soybean.

Cadmium (Cd) is a hazardous heavy metal pollutant that poses significant risks to agricultural production and human health. Nanoparticles (NPs) can alleviate the effects of cadmium on crops by regulating the expression of stress-responsive genes, however, the mechanism of regulation is unknown. N6-methyladenosine (m6A) is a prevalent RNA modification, which determines the expression level of RNA. In this study, we performed m6A methylome analysis and gene editing to investigate the regulation of stress-responsive genes by m6A under Cd stress with ZnO nanoparticles (ZnO NPs). Firstly, we identified 16 differentially expressed genes (DEGs) with differential m6A-modification by m6A methylome seq, which included 6 stress-responsive genes. ZnO NPs treatment reduced the m6A level and stabilized the mRNAs of these stress-responsive genes. Then, we utilized the in vivo m6A modification tool, Plant m6A Editors (PMEs), specifically reduced the m6A level of NRT1 and GM35E in transgenic plants. The expression of NRT1 and GM35E was increased, and plants carrying PMEs-NRT1 and PMEs-GM35E showed stronger Cd tolerance than control. Therefore, we can conclude that NPs enhance Cd tolerance in plants by regulating the m6A methylation level of stress-responsive genes such as NRT1 and GM35E, which ultimately affects the expression of these genes. This study proposed a post-transcriptional regulatory network in vegetable soybean roots responding to Cd with ZnO NPs, potentially offering new insights into gene manipulation for controlling low Cd accumulation in crops.