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Jochen HM Prehn
Caoimhin G Concannon
Slavomir Kacmar
Heiko Duessmann
Markus Rehm
Manus W Ward



physics recombinant fusion proteins neurons receptors glutamate cells cultured tumor necrosis factor cells caspases receptors tumor necrosis factor membrane potentials humans physiology rats bh3 interacting domain death agonist protein mitochondria cultured glutamate glutamic acid animals fluorescent dyes receptors fluorescence resonance energy transfer apoptosis

Real time single cell analysis of Bid cleavage and Bid translocation during caspase-dependent and neuronal caspase-independent apoptosis. (2006)

Abstract Bcl-2 homology domain (BH) 3-only proteins couple stress signals to evolutionarily conserved mitochondrial apoptotic pathways. Caspase 8-mediated cleavage of the BH3-only protein Bid into a truncated protein (tBid) and subsequent translocation of tBid to mitochondria has been implicated in death receptor signaling. We utilized a recombinant fluorescence resonance energy transfer (FRET) Bid probe to determine the kinetics of Bid cleavage and tBid translocation during death receptor-induced apoptosis in caspase 3-deficient MCF-7 cells. Cells treated with tumor necrosis factor-alpha (200 ng/ml) showed a rapid cleavage of the Bid-FRET probe occurring 75.4 +/- 12.6 min after onset of the tumor necrosis factor-alpha exposure. Cleavage of the Bid-FRET probe coincided with a translocation of tBid to the mitochondria and a collapse of the mitochondrial membrane potential (DeltaPsim). We next investigated the role of Bid cleavage in a model of caspase-independent, glutamate-induced excitotoxic apoptosis. Rat cerebellar granule neurons were transfected with the Bid-FRET probe and exposed to glutamate for 5 min. In contrast to death receptor-induced apoptosis, neurons showed a translocation of full-length Bid to the mitochondria. This translocation occurred 5.6 +/- 1.7 h after the termination of the glutamate exposure and was also paralleled with a collapse of the DeltaPsim. Proteolytic cleavage of the FRET probe also occurred, however, only 25.2 +/- 3.5 min after its translocation to the mitochondria. Subfractionation experiments confirmed a translocation of full-length Bid from the cytosolic to the mitochondrial fraction during excitotoxic apoptosis. Our data demonstrate that both tBid and full-length Bid have the capacity to translocate to mitochondria during apoptosis.
Collections Ireland -> Royal College of Surgeons in Ireland -> Physiology and Medical Physics Articles
Ireland -> Royal College of Surgeons in Ireland -> Department of Physiology and Medical Physics

Full list of authors on original publication

Jochen HM Prehn, Caoimhin G Concannon, Slavomir Kacmar, Heiko Duessmann, Markus Rehm, Manus W Ward

Experts in our system

Jochen H M Prehn
Royal College of Surgeons in Ireland
Total Publications: 206
CaoimhĂ­n G Concannon
Royal College of Surgeons in Ireland
Total Publications: 46
Heiko Duessmann
Royal College of Surgeons in Ireland
Total Publications: 5
Markus Rehm
Royal College of Surgeons in Ireland
Total Publications: 55