|
|
|
|
|
|
|
||
J. Cell Biol.,
Volume 144, Number 5, March 8, 1999 903-914
Cancer Research Campaign Molecular and Cellular Pharmacology Group, School of Biological Sciences, University of
Manchester, Manchester M13 9PT, United Kingdom
Investigation of events committing cells to
death revealed that a concealed NH2-terminal epitope
of the pro-apoptotic protein Bak became exposed in
vivo before apoptosis. This occurred after treatment of
human Jurkat or CEM-C7A T-lymphoma cells with the mechanistically disparate agents staurosporine, etoposide or dexamethasone. The rapid, up to 10-fold increase in Bak-associated immunofluorescence was
measured with epitope-specific monoclonal antibodies
using flow cytometry and microscopy. In contrast, using
a polyclonal antibody to Bak, immunofluorescence was
detected both before and after treatment. There were
no differences in Bak protein content nor in subcellular
location before or after treatment. Immunofluorescence showed Bcl-xL and Bak were largely associated
with mitochondria and in untreated cells they coimmunoprecipitated in the presence of nonioinic detergent.
This association was significantly decreased after cell
perturbation suggesting that Bcl-xL dissociation from Bak occurred on exposure of Bak's NH2 terminus. Multiple forms of Bak protein were observed by two dimensional electrophoresis but these were unchanged
by inducers of apoptosis. This indicated that integration
of cellular damage signals did not take place directly on
the Bak protein. Release of proteins, including Bcl-xL, from Bak is suggested to be an important event in commitment to death.
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?
This article has been cited by other articles:
|
|
