SiRAP2-12, a Positive Regulatory Factor, Effectively Improves the Waterlogging Tolerance of Foxtail Millet (Setaria italica)

Foxtail millet (Setaria italica) growth was inhibited because of waterlogging stress, which has caused yield reduction. ERF family plays an important role to plant adversity tolerance. In our study, we obtained 19,819 differential expressed genes (DEGs) between the two treatments based on the RNA-seq sequencing of foxtail millet of waterlogging stress. Furthermore, a total of 28 ERF family members were obtained, which have a complete open reading frame. We studied the evolution and function of SiERF family and how they affected the waterlogging tolerance. It was found that SiERF1A/B/C (GenBank ID: OR775217, OR775219, OR775218) and SiRAP2-12 (GenBank ID: OR775216) have similar functions to the known waterlogging tolerance genes of other plants. Among them, the SiRAP2-12 expression was obviously significantly up-regulated in foxtail millet after 5d waterlogging stress. After SiRAP2-12 was silenced, the activity of defense enzymes in millet decreased significantly. In details, superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD), the osmotic regulator proline (Pro), and the activity of the anaerobic respiratory enzyme alcohol dehydrogenase (ADH) content were decreased by 78.61%, 29.52%, 79.95%, 19.41% and 54.77%, respectively. In contrast, the relative electrical conductivity contents (REC), malondialdehyde (MDA), and hydrogen peroxide (H₂O₂) of the foxtail millet subjected to virus-induced gene silencing clearly increased by 1.03-fold, 36.09%, and 15.21%, respectively. The content of sodium (Na⁺) in the SiRAP2-12-silenced foxtail millet also increased, but that of potassium (K⁺) decreased. Interestingly, we found that ethylene content was significantly reduced. Further, the SiAOC1 expression, an essential gene for ethylene synthesis, was inhibited in SiRAP2-12-silenced foxtail millet after waterlogging stress. Taken together, we hypothesized that SiRAP2-12 might be a positive regulator of millet tolerance to waterlogging stress.