Bcl-2 mediated inhibition of erucylphosphocholine-induced apoptosis depends on its subcellular localisation.
The synthetic phospholipid derivative erucylphosphocholine (ErPC) is a potent inducer of apoptosis in human tumor cell lines. This membrane-targeted drug induces apoptosis independently from death receptor signaling through a mitochondrial pathway that is inhibited by over-expression of Bcl-2. Within the cell, Bcl-2 resides in membranes of mitochondria, endoplasmic reticulum (ER) and the nucleus. However, the importance of its subcellular localisation in distinct organelles for protection against apoptosis is not completely understood. To investigate the impact of Bcl-2 localised at defined subcellular compartments on its protective effects against ErPC-induced apoptosis, Bcl-2 expression was directed to the outer membrane of the mitochondria or the ER of Jurkat T Lymphoma cells, using Bcl-2 mutants with modified membrane anchors. The mitochondrial insertion sequence of ActA directed Bcl-2 to the mitochondria (Bcl-2/MT), the ER-specific sequence of cytochrome b5 to the ER (Bcl-2/ER). Additionally, Jurkat cells expressing wild-type Bcl-2 (Bcl-2/WT) or a transmembrane domain-lacking mutant (Bcl-2/DeltaTM) were employed. While restricted expression of Bcl-2 either at membranes of the mitochondria or the ER strongly interfered with ErPC-induced mitochondrial damage and apoptosis, cytosolic Bcl-2/DeltaTM exhibited only reduced protection. Thus, membrane localisation of Bcl-2 is a prerequisite for substantial protection against ErPC-induced apoptosis. For efficient long-term inhibition of ErPC-induced apoptosis Bcl-2 had to be present in the membranes of both compartments, the ER and the mitochondria. The finding that ER-targeted Bcl-2 interferes with ErPC-induced mitochondrial damage points to an involvement of the ER in apoptosis signaling upstream of the mitochondria and to a crosstalk between both compartments.
Handrick
R
R
Department of Radiation Oncology, University of Tübingen, Hoppe-Seyler-Strasse 3, 72076 Tübingen, Germany.
Rudner
J
J
Müller
I
I
Eibl
H
H
Belka
C
C
Jendrossek
V
V
D017209:Apoptosis / Q000187:drug effects*; D020169:Caspases / Q000378:metabolism; D045304:Cytochromes c / Q000378:metabolism; D004789:Enzyme Activation; D006801:Humans; D007425:Intracellular Membranes / Q000502:physiology; D019169:Jurkat Cells; D008564:Membrane Potentials; D008928:Mitochondria / Q000502:physiology; D010767:Phosphorylcholine / Q000031:analogs & derivatives* / Q000494:pharmacology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000502:physiology*; D013347:Subcellular Fractions / Q000378:metabolism*
D019253:Proto-Oncogene Proteins c-bcl-2; D010767:Phosphorylcholine; C075010:erucylphosphocholine; D045304:Cytochromes c; D020169:Caspases