There is an error in panel D of S8 Fig. Specifically, the upper panel should read ‘SclB-cYFP + nYFP’, not ‘SclA-cYFP + nYFP’. The authors have provided a corrected version here.
GFP-fusion proteins of SclB are functional and phosphorylated and Bi-FC controls are negative.
A) Strains expressing SclB either N- or C-terminally tagged with sGFP in ΔsclB background, ΔsclB and wildtype (WT) were point inoculated on solid MM and grown for 4 days in light. B) SclB-GFP and GFP-SclB fusion proteins expressed under native promoter are visualized in a western hybridization assay employing an α-GFP antibody (GFP) and Ponceau staining as loading control (Pnc). The black arrow indicates bands corresponding to full-length fusion proteins (in silico prediction 87.46 kDa). C) Protein crude extracts of GFP-SclB grown vegetatively were mixed with phosphatase inhibitor cocktail (-/PhoI), with Lambda phosphatase (λ/-), or Lambda phosphatase and phosphatase inhibitor cocktail (λ/PhoI). A control sample was left untreated (-/-). A subsequent western hybridization assay employing α-GFP antibody visualizes protein bands. D) Two strains, either expressing sclB::cyfp and the free second half of the split YFP (nyfp; upper part), or free cyfp and rcoA::nyfp (lower part), under control of a bi-directional nitrate promoter were constructed. Strains were inoculated in liquid MM and analyzed with fluorescence microscopy after 36 h at 30°C.(TIF)Click here for additional data file.
Authors: Karl G Thieme; Jennifer Gerke; Christoph Sasse; Oliver Valerius; Sabine Thieme; Razieh Karimi; Antje K Heinrich; Florian Finkernagel; Kristina Smith; Helge B Bode; Michael Freitag; Arthur F J Ram; Gerhard H Braus Journal: PLoS Genet Date: 2018-07-25 Impact factor: 5.917