Over-expression of GAPDH induces apoptosis in COS-7 cells transfected with cloned GAPDH cDNAs

Abstract

The cDNAs for glyceraldehyde-3-phosphate dehydrogenase (GAPDH; EC 1.2.1.12) were cloned from cerebellar neurons undergoing age-induced apoptosis and/or healthy cells. COS-7 cells were transfected with the isolated GAPDH cDNAs using to the Lipofectamine method. Assessment of cell death in this paradigm was performed by monitoring the co-transfected luciferase activities and the characterization of cell death was examined by the DNA fragmentation assay and Hoechst dye nuclear staining. These observations show that over-expression of GAPDH occurring from both cDNAs robustly induces apoptotic death in the transfected COS-7 cell cultures. Confocal-immunocytochemical studies using this GAPDH-specific monoclonal antibody revealed that nuclear translocation of overexpressed GAPDH is a primary apoptotic event. Our results directly demonstrate that over-expressed GAPDH functions as a 'killing protein' in apoptosis.