Fig. 6. Examination of subcellular localization and protein-protein interaction in living cells for de novo-expressed Src and TRAF6. Bimolecular fluorescence complementation (BiFC) involving split YFP tags was used to detect intracellular protein-protein interaction between TRAF6 and Src that could be compared with the expression pattern for these proteins tagged with full-length YFP in HEK293T cells transiently transfected for 24 hours. (A) Transfection of 200 ng of individual, uncomplemented BiFC vectors; –YN, N-terminal YFP fragment (aa 1-154), –YC, C-terminal YFP fragment (aa 155-238). (B) Transfection of 200 ng of each BiFC vector complementary pair (–YN + –YC). (C) Subcellular localization of Src-YFP and variants in cells transfected with 50 ng of expression vectors encoding each protein form, as indicated. (D) Subcellular localization of TRAF6-YFP and variants in cells transfected with 20 ng of expression vectors as indicated. (E) Image enhancement of Fig. 6B panels a and c, revealing distinct TRAF6 and Src morphologies.