The role of CsaMIR396E-CsaGRFs in regulating root:shoot ratio under osmotic stress in cucumber

As sessile organisms, plants can modify their growth to adapt to the changed environment. Here, our results showed that shoot growth was more severely inhibited than root growth, resulting root:shoot ratio was increased in cucumber under osmotic stress. CsaMIR396E was highly expressed under well-watered condition, however, it was dramatically down-regulated to release its Growth-regulating factor (GRF) targets under osmotic stress in roots, but not in shoots. Exogenous abscisic acid (ABA) treatment suggested that down-regulation of CsaMIR396E in roots was depended on ABA signal. CmoMIR396E, the homolog of CsaMIR396E in pumpkin, exhibited a similar osmotic stress response pattern in pumpkin roots, and bioinformatic analysis showed that two motifs were conservatively presented within the promoter of MIR396E in cucurbits. Motif 1 harbored an ABA-response element (ABRE), while motif 2 harbored a (CT)(n)/(GA)(n )dinucleotide repeat element and functioned as an enhancer. Additionally, the role of CsaMIR396E in regulating root:shoot ratio under osmotic stress was confirmed by transgenic overexpression in Arabidopsis. In summary, our results suggested that CsaMIR396E acts as an osmotic stress response gene in roots, and it regulates root:shoot ratio by miR396-GRFs pathway under osmotic stress in cucumber.