Antisense BAG-1 sensitizes HeLa cells to apoptosis by multiple pathways.

To study the mechanism of action of BAG-1 in drug-induced apoptosis, we constructed an antisense BAG-1 vector and established a stably transfected cell line from BAG-1-over-expressing HeLa cells. Reduced BAG-1 protein was confirmed by Western blot. Treatment of the antisense BAG-1-transfected cells with the anti-cancer drugs staurosporine, paclitaxel, all-trans retinoic acid (ATRA), and N-(4-hydroxyphenyl) retinamide (4-HPR) resulted in significantly enhanced apoptosis and reduced cell viability relative to vector-transfected cells. While the expression of p53 was increased, the level of Bcl-2 and Bax was decreased. Cells underexpressing BAG-1 had reduced cytosolic cytochrome c level. Treatment with staurosporine and paclitaxel resulted in increased cytochrome c release from mitochondria, whereas there was no change induced by treatment with ATRA and 4-HPR. Our experiments suggest that BAG-1 inhibits anti-cancer drug-induced apoptosis through apoptosis regulation pathways that may involve the mitochondrial Bcl-2/Bax ratio, p53, and differential anti-cancer drug-mediated cytochrome c release.