Overexpression of bad preferentially augments anoikis

BAD is a BH3-only protein, and its proapoptotic activity is negatively regulated by serine phosphorylation. Here, we show that overexpression of BAD preferentially augments anchorage loss-induced apoptosis (anoikis). Gene transfer-mediated BAD overexpression alone did not induce apoptosis in attached MDCK cells but strongly augmented apoptosis when cells were cultured in suspension. In contrast, overexpression of another BH3-only protein, BID, displayed much lower augmentation of anoikis, suggesting a preferential contribution of BAD to anoikis. During suspension culture, un-phosphorylated BAD was gradually increased and targeted to the mitochondria. Cotransfection of BAD with constitutively active Akt cDNA strongly inhibited this change. In contrast, the increase of un-phosphorylated BAD was not significantly inhibited by several phosphatase inhibitors or cotransfection with a dominant negative calcineurin cDNA, implying that the increase may be mainly due to a decrease of serine kinase activity, such as that of Akt. Similar results were observed in COS-7 cells, suggesting that BAD overexpression can increase sensitivity of anchorage-dependent cancer cells to anoikis. Thus, we propose that BAD can serve as a valuable gene therapeutic molecule to inhibit carcinoma progression. (C) 2003 Wiley-Liss, Inc.