Fig. 5: Inhibition of chlorophyll degradation in SlZHD17-RNAi fruit by directly regulating SlSGR1.
From: SlZHD17 is involved in the control of chlorophyll and carotenoid metabolism in tomato fruit
a Photograph of the cross section of WT and SlZHD17-RNAi fruit at the Br+3 stage. Bar = 5 mm. b, c The contents of chlorophyll a (b) and chlorophyll b (c) in WT and SlZHD17-RNAi fruit at different stages detected by high-performance liquid chromatography (HPLC). FW fresh weight, ND not detected. d The relative expression level of SlSGR1 in WT and SlZHD17-RNAi Br+3 fruit. For each transgenic fruit, the normal orange part (Or) and abnormal green part (Gr) were sampled separately. In b–d, data represent the mean values of three independent experiments, and error bars show the standard error values. Single asterisk (*) and double asterisks (**) refer to significant differences between WT and transgenic lines with P < 0.05 and P < 0.01, respectively (two-tailed Student’s t-test). e SlZHD17 activates the SlSGR1 gene promoter, as determined by dual-luciferase assay. The empty effector was used as a control (set as 1). Data represent the mean values of six independent experiments, and error bars show the standard error values. Double asterisks (**) refer to significant difference between empty effector and SlZHD17-effector with P < 0.01 (two-tailed Student’s t-test). The P value is provided in the graph. f Schematic diagram of the sequence positions chosen for yeast one-hybrid assay and electrophoretic mobility shift assay (EMSA). The underlined region indicates the promoter fragment used for the yeast one-hybrid assay, and the triangle indicates the sequence position used for the EMSA. The detailed sequences are provided in Appendix S4. g SlZHD17 binding with the SlSGR1 promoter fragment assessed by yeast one-hybrid assay. The yeast transformants were cultured on SD/−Leu and SD/−Leu/+AbA media for 3–5 days, and the pGADT7-empty plasmid was used as a control. h SlZHD17 binding to the SlSGR1 promoter region in vitro by EMSA. TF-His protein incubated with biotin-labeled DNA probe was used as a negative control. ‘+++’ indicates an increase in the competitor probe; no shift in the band for the His-SlZHD17 protein and mutant biotin-labeled DNA probe further confirmed the specific binding site
