BCL-2 MUTANTS WITH RESTRICTED SUBCELLULAR LOCATION REVEAL SPATIALLY DISTINCT PATHWAYS FOR APOPTOSIS IN DIFFERENT CELL-TYPES

Human Bcl-2 is located in multiple intracellular membranes when expres sed in MDCK and Rat-1/myc cells. We restricted expression to the endop lasmic reticulum or mitochondria by exchanging the Bcl-2 carboxytermin al insertion sequence for an equivalent sequence from cytochrome b5 or ActA, respectively MDCK cells are protected from serum deprivation-in duced apoptosis by both wild-type Bcl-2 and the mutant targeted to mit ochondria but not by the mutant targeted to endoplasmic reticulum. In contrast, when expressed in Rat-1/myc cells, the Bcl-2 mutant located at the endoplasmic reticulum is more effective than that targeted to m itochondria, In MDCK cells both mutants bind Bax as effectively as wil d-type, demonstrating that Bax binding is not sufficient to prevent ap optosis.