Observation of interaction between bid and 14-3-3 proteins by FRET in living cell during TNF-α-induced apoptosis

Caspase8 is activated and cleaves Bid into two fragments when cells are exposed to death-inducing molecules such as tumor necrosis factor-alpha(TNF- alpha). Then the C-terminal fragment relocates from cytosol to mitochondria and promotes the release of cytochrome c, in the final cellular apoptosis is induced. Despite recent progress in the study of Bid during apoptosis induction, it remains unclear how C-terminal fragment of Bid cleaved moves to mitochondria and then induces the release of cytochrome c and so on. The 14-3-3 proteins are known to sequester certain pro-apoptotic members of Bcl-2 family. In order to further study the biological action of Bid during apoptosis, especially under physiological condition of living cell, the plasmids pBid-CFP and pYFP- 14-3-3 were constructed. By the transient transfection of pBid-CFP and pYFP-14-3-3, the dynamic process of interaction of Bid and 14-3-3 protein in individual living cell during the apoptosis was primarily investigated with FRET(fluorescent resonance energy transfer) technique by the use of fluorescence microscopy.