Alzheimer's disease, a progressive neurodegenerative disorder, is
characterized by deposition of Aβ, accumulation of intercellular
neurofibrillary tangles, and neuronal cell loss. Recently, it was
reported that the activation of caspase 8 was observed in Aβ-induced
neuronal apoptosis. In general, when cells are exposed to death-inducing
molecules such as TNF-α or Fas, caspase 8 is activated and cleaves
an apoptotic facilitator Bid that is a member of the Bcl-2 family.
After additional modification, the carboxy-terminal moiety of Bid
is translocated to the mitochondria and induces the release of cytochrome
c into the cytoplasm. In an attempt to directly observe the cleavage
of Bid and the following events in living cells, we constructed a
vector that encoded Bid fused with YFP and CFP (YFP-Bid-CFP). Upon
expression of YFP-Bid-CFP in mammalian cells, we were able to observe
the efficient transfer of energy from excited CFP to YFP within the
YFP-Bid-CFP molecule, and, importantly, the fusion protein YFP-Bid-CFP
was fully functional in cells. When YFP-Bid-CFP was cleaved by caspase
8, upon activation by anti-Fas antibodies but not by Aβ or tunicamycin,
no such transfer of energy was detected. This data suggests that other
apoptotic pathways are more important in Ab-induced apoptosis.
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| Activation of caspase 8 was
detected in single cells by disappearance of Fluorescence energy
resonance transfer (FRET). CFP-Bid-YFP-expressed COS 7 cells
were incubated with TNFa and cycloheximide (CHX) to induce cell
death. In general, caspase 8 is activated by TNF-α
and CHX. |
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| Relational Information |
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AIST Today Vol. 3, No. 3 (2003) 11
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No.8
Spring 2003
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