Fig. 6: Transcription factor NtCYC rewrites aroma formation.

a The sub-network extracted from the GMRN for carotenoid degradation. SG, structural genes. TF, transcription factors. b Expression patterns of metabolites, SGs, and TFs. “4” and “-4” represent the maximum and minimum value of the Z-score standardized expression, respectively. DAT, days after topping. c Transcriptional activation assay of NtCYC in yeast. d Subcellular localization of NtCYC-GFP fusion protein in Nicotiana benthamiana leaf cells. GFP, green fluorescent protein. mKATE, Katushka protein. Scale bars, 20 µm. e Transient expression assay showing transcriptional activation of the LUC reporter gene (driven by the promoter of NtLOX2) by NtCYC (n = 5 biological replicates). *** P < 0.001. f Specific binding of NtCYC protein to promoters of NtLOX2 by one-hybrid system using bait and either prey or negative control (bait/pGADT7). g EMSA with NtCYC protein performed with the probes derived from the NtLOX2 promoter. h–j Dihydroactinidiolide (h), damascenone (i) and ionone (j) contents and corresponding mass spectrometry data in leaves of WT, NtCYC-OE and ntcyc-KO tobaccos (n = 3 biological replicates). k RNA-seq and RT-qPCR showing that NtLOX2 are positively regulated by NtCYC (n = 3 biological replicates). l MDA content between WT, NtCYC-OE and ntcyc-KO tobaccos (n = 3 biological replicates). m GMRN accurately predicts targets of NtCYC. The P values were calculated using hypergeometric distribution (two-sided) and then adjusted using the FDR. n Expression patterns of the genes related to carotenoid pathway in WT, NtCYC-OE and ntcyc-KO tobaccos. “2” and “−2” represent the maximum and minimum value of the Z-score standardized expression, respectively. In c, d, f, and (g), the images were representative of three independent repeats with similar results. In (e), (h), (i), (j), (k), and (l), error bars represent the means ± standard errors of biological replicate. A two-tailed Student’s t test was used to determine P values. Different lowercase letters indicate significant differences (P < 0.001). Source data are provided as a Source Data file.