The Rice <i>Cis</i> ‐Natural Antisense Transcript <scp> <i>NAT1850</i> </scp> of <i>Pri‐</i> <scp> <i>miR1850</i> </scp> Negatively Regulates Cold Tolerance by Repressing <scp> <i>NPR3</i> </scp>

Natural antisense transcripts (NATs) correspond to nearly 60% of annotated rice loci, however their functions are largely unknown. In this study, we characterise a rice cis-NAT (NAT1850) that completely overlaps with a rice-specific primary miRNA, pri-miR1850. Pri-miR1850, but not its mature miR1850 products, promotes the accumulation of NAT1850, while NAT1850 overexpression in turn reduces the accumulation of pri-miR1850 transcripts. A 21-nt siRNA (siR1850) derived from the pri-miR1850 transcripts is generated by cleavage of pri-miR1850-NAT1850 dsRNA and overlaps in sequence with miR1850.1 and miR1850.2. Both NAT1850 and siR1850 negatively regulate cold tolerance at both the young-seedling and booting stages. Interestingly, siR1850 targets and represses NPR3, which is also a target of miR1850.1. NPR3 interacts with the WRKY76 transcription factor and acts as a co-transcriptional activator of WRKY76 to trigger DREB1B under cold stress. Genetic evidence shows the NAT1850-siR1850 module functions in cold stress response via an NPR3-dependent manner. Furthermore, NAT1850 and siR1850 control nitrogen assimilation and rice yield in a miR1850.1-NPR3-independent pathway. Our findings reveal a regulatory mode for a pri-miRNA and its cis-NAT, and uncover their roles in balancing the cold-stress response and rice yields.