Regulation of a Novel Carbon Monoxide-Responsive miRNA in Sweet Potato

Carbon monoxide (CO), a gaseous secondary messenger, plays a crucial role in regulating growth and stress responses in plants. MicroRNAs (miRNAs), which are small noncoding RNAs comprising 21-24 nucleotides, modulate gene expression by directing messenger RNA cleavage or inhibiting translation. However, the impact of CO on miRNAs is rarely investigated in plants. In this study, we examined the regulatory interactions between CO and miRNAs in sweet potato (Ipomoea batatas 'Tainung 57'). A small RNA library prepared from CO-treated sweet potato leaves was constructed and analyzed using next-generation sequencing technology. A CO-induced novel 22-nucleotide-long miRNA, designated tag202, was identified. The expression of tag202 was increased in response to CO treatment. Overexpression and knockdown studies of tag202 in transgenic sweet potato revealed an inverse relationship between tag202 expression and its predicted target gene, which encodes a Malectin-like domain-containing receptor-like kinase (IbMLD-RLK) localized to the plasma membrane. The IbMLD-RLK mRNAs were predominantly cleaved at the canonical cleavage position within the tag202 binding region, which was mediated by tag202. The IbMLD-RLK protein was indicated to interact with IbMEK1, a component of the mitogen-activated protein kinase (MAPK) signaling cascade. In response to wounding, the expression of pri-tag202 and IbMLD-RLK was decreased and increased, respectively. Exogenous CO supplementation disrupted this response. In conclusion, CO-induced tag202 acts as a regulatory switch, targeting IbMLD-RLK to modulate the MAPK signaling pathway. This study offers new insights into the molecular mechanisms of CO-responsive miRNAs and their roles in regulating responses to wounding.