FIGURE 1. Yeast three-hybrid screen to identify Fyn phosphorylation-dependent NR2B interactants.
Yeast were co-transformed with a rat brain cDNA library and a bait plasmid that expressed the cytoplasmic tail of the NR2B subunit of the NMDAR (ctNR2B) in-frame with a Gal4 DNA binding domain (BD) and a constitutively active form of Fyn (FynCA). FynCA expression was governed by a conditional methionine promoter, such that FynCA expression was induced in the absence of methionine (−met) (A) and repressed in the presence of methionine (+met) (B) in the yeast growth media. Colonies turning blue in the absence of methionine (C), but not in the presence of methionine (D), were identified as potential three-hybrid interactants reliant on the expression of FynCA.