3.5 CRIR1 regulates cold stress-related genes in a CBF-independent pathway
We next deployed the genes falling into the category of response to the abiotic stimulus, which included two transcription factors (MeNFYA and MeNAC ), GALACTINOL SYNTHASE(MeGOLS ), and ABSCISIC STRESS-RIPENING PROTEIN(MeASR ). We verified the expression level of these genes in OE lines with qRT-PCR (Figure 5a). Consistently, transcription levels ofMeNFYA and MeNAC were increased in OE lines compared to WT plants under both normal and cold-treated conditions. Quantification of the MeGOLS showed similar trends, whereas we could only detectMeASR after cold stress, suggesting that MeASR is accumulated under low temperature more than under normal conditions. In our RNA-seq analysis, we also found that several IAA-signaling genes were expressed at a lower level in cold-treated OE seedlings than in WT seedlings. These genes include transcription factor YABBY2 ,AUXIN-RESPONSIVE PROTEIN (IAA18 ), and AUXIN-BINDING PROTEIN (ABP19 ). Real-time PCR analysis showed a consistent result with transcriptomic data (Figure 5b). Additionally, we examined the expression patterns of classical cold stress markers, includingMeCBFs , MeCORs , and MeICE1 . Accumulated evidence has revealed that CBF proteins function as key regulators of the transcriptomic rearrangement leading to cold acclimation. In this study, the WT seedlings showed increased MeCBFs  and MeCOR  expression levels under cold stress. However, the expression levels of these genes induced by cold stress in the OE seedlings were similar to those of the wild type (Figure 5c). In conclusion, these results indicate that modulation of CRIR1 significantly regulates cold stress-related genes in a CBF -independent pathway.