title: Enhanced heterodimerization of Bax by Bcl-2 mutants improves irradiated cell survival.
abstract: B Cell Lymphoma-2 (Bcl-2) protein suppresses ionizing radiation-induced apoptosis in hemato-lymphoid system. To enhance the survival of irradiated cells, we have compared the effects and mechanism of Bcl-2 and its functional variants, D34A (caspase-3 resistant) and S70E (mimics phosphorylation on S70). Bcl-2 and its mutants were transfected into hematopoietic cell line and assessed for cell survival, clonogenicity and cell cycle perturbations upon exposure to ionizing radiation. The electrostatic potential of BH3 cleft of Bcl-2/mutants and their heterodimerization with Bcl-2 associated X protein (Bax) were computationally evaluated. Correspondingly, these results were verified by co-immunoprecipitation and western blotting. The mutants afford higher radioprotective effect than Bcl-2 in apoptotic and clonogenic assays at D(0) (radiation dose at which 37 % cell survival was observed). The computational and functional analysis indicates that mutants have higher propensity to neutralize Bax protein by heterodimerization and have increased caspase-9 suppression capability, which is responsible for enhanced survival. This study implies potential of Bcl-2 mutants or their chemical/peptide mimics to elicit radioprotective effect in cells exposed to radiation.
authors.affiliation: Stem Cell and Gene Therapy Research Group, Institute of Nuclear Medicine and Allied Sciences (INMAS), Lucknow Road, Timarpur, Delhi 110054, India.
authors.forename: Yogesh Kumar
authors.initials: YK
authors.lastname: Verma
authors.forename: Pawan Kumar
authors.initials: PK
authors.lastname: Raghav
authors.forename: Hanumantharao Guru
authors.initials: HG
authors.lastname: Raj
authors.forename: Rajendra Prasad
authors.initials: RP
authors.lastname: Tripathi
authors.forename: Gurudutta U
authors.initials: GU
authors.lastname: Gangenahalli
mesh_terms: D017209:Apoptosis / Q000528:radiation effects*; D053453:Caspase 9 / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival; D006801:Humans; D008958:Models, Molecular; D055503:Protein Multimerization; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000737:chemistry* / Q000378:metabolism
chemical_list: D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D053453:Caspase 9