Fig. 3: Identification of the functional phosphorylation sites for FTZ-F1.

a Immunoprecipitation of exogenous FTZ-F1 from Sf9 cells using anti-GFP. b The putative phosphorylation site T288 of FTZ-F1 was identified by LC-MS/MS. The precursor ion, which ranges from 264 to 302, is shown in the top band with the amino acids in white letters, and the representative fragment ion is shown with a red background with detailed information. The identified phosphorylation site T288 is represented by a red letter and subscripted p, while the phosphorylated peptide on the spectrum is underlined in red. c The DEPP score of the four putative phosphorylation sites was computed using the DEPP software in silico. d The regulatory effect of phosphorylated FTZ-F1^T288D(P) and non-phosphorylated FTZ-F1^T288A(-P) on midgut genes. A substitutes T mimicking dephosphorylation and D substitutes T mimicking sustained phosphorylation. e, f WebLogo plots highlight the potential functional FBS in receptor gene promoters (e) and FBS^P in non-receptor gene promoters (f). g Effect of non-phosphorylated FTZ-F1^T288A(-P) on the activity of receptor promoters with either a wild-type or a mutant FBS. For clarity, only the functional FBSs are shown. h Effect of phosphorylated FTZ-F1^T288D(P) on the activity of non-receptor promoters with either a wild-type or a mutant FBS^P. The empty pAc5.1 vector was used as a control (d, g, h), and the relative luciferase activity (fold) was calculated based on the value of the control, which was assigned an arbitrary value of 1. Data were presented as mean values ± SEM (n = 3), ns, not significant, p values are shown. One-way ANOVA with Tukey’s test was used for comparison. Source data are provided as a Source Data file.