Fig. 5: FgSfp1 interacts with FgSch9 and FgMaf1 as a downstream TF of the nutrient stress sensing FgTOR signaling pathway for regulating mycotoxin biosynthesis.

a Y2H assay determined that FgSfp1 interacts with and FgSch9. Yeast cells co-expressing FgSch9-pGADT7 and FgSfp1-pGBKT7 plasmids were plated on selective plates. Experiments were conducted successfully three times independently. b Bi-FC assay confirms that FgSfp1 interacted with FgSch9 in F. graminearum by YFP fluorescence detection with a confocal laser scanning microscope. Wild-type PH-1 co-expressing FgSfp1-NYFP and FgSch9-CYFP fusion proteins were incubated in YEPD medium for 48 h and observed using a confocal laser scanning microscope. The fungal nucleus was simultaneously stained by DAPI. DIC, differential interference contrast. Scale bar = 10 μm. c Y2H assay determined that FgSfp1 interacts with and FgMaf1. Yeast cells co-expressing FgMaf1-pGADT7 and FgSfp1-pGBKT7 plasmids were plated on selective plates. Experiments were repeated three times independently. d Rapamycin-treatment assay demonstrates FgTOR signaling pathway inhibition leads to FgSfp1 retaining at cytoplasm observed with fluorescent microscope. Conidia of FgSfp1-GFP overexpression strain were used for 12 h rapamycin treatment. Final concentration at 120 µg/mL, same volume of solvent was added as mock group. Experiments were repeated for three times independently. e Different colony morphology of wild-type PH-1 and ∆FgSfp1 mutant strains on complete medium (CM) supplemented with a gradient concentration of rapamycin incubated at 28 °C for 4 days. f Inhibition rate of rapamycin on PH-1 and ∆FgSfp1 calculated from colonies diameter measurements from (b). The P-values were evaluated using t-test, *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001.