BAX HOMODIMERIZATION IS NOT REQUIRED FOR BAX TO ACCELERATE CHEMOTHERAPY-INDUCED CELL-DEATH

Bax, a member of the Bcl-2 family of proteins, has been shown to accel erate apoptosis induced by growth factor withdrawal, gamma-irradiation , and the chemotherapeutic agent, etoposide, The mechanism by which Ba x promotes apoptosis is poorly understood, Bax forms homodimers which have been suggested to act as accelerators or inducers of cell death, However, the requirement for homodimerization of Bax to promote cell d eath remains unclear. We performed site-directed mutagenesis of the BH 1, BH2, and BH3 in Bax to determine the regions of Bar required for ho modimerization and to define the role of fax homodimers in cell death induced by chemotherapy drugs, Bar proteins expressing alanine substit utions of the highly conserved amino acids glycine 108 (G108) in BH1, tryptophan 158 (W158) in BH2, and glycine 67 and aspartic acid 68 (GD6 7-68) in BH3 as well as deletion of the most conserved amino acids in BH1 (Delta 102-112) and BH2 (Delta 151-159) and deletion of BH3 (Delta 63-71) maintained their ability to accelerate chemotherapy-induced ce ll death, Immunoprecipitation studies revealed that Bax with deletions in BH1 and BH2 still associated with wild-type Bax while deletion of BH3 disrupted Bar homodimerization. These results demonstrate that Bar does not require the conserved regions of homology, BH1, BH2, or BH3, to accelerate chemotherapy-induced cell death, Furthermore, our resul ts established BH3 as a region required for Bax homodimerization in ma mmalian cells and demonstrate that monomeric forms of Bar are active i n accelerating cell death induced by chemotherapy agents.