Bid binding to negatively charged phospholipids may not be required for its pro-apoptotic activity in vivo

Bid is a ubiquitous pro-apoptotic member of the Bcl-2 family that has been involved in a variety of pathwaysof cell death. Unique among pro-apoptotic proteins, Bid is activated after cleavage by the apical caspases ofthe extrinsic pathway; subsequently it moves to mitochondria, where it promotes the release of apoptogenicproteins in concert with other Bcl-2 family proteins like Bak. Diverse factors appear to modulate the proapoptoticaction of Bid, from its avid binding to mitochondrial lipids (in particular, cardiolipin) to multiplephosphorylations at sites that can modulate its caspase cleavage. This work addresses the question of howthe lipid interactions of Bid that are evident in vitro actually impact on its pro-apoptotic action within cells.Using site-directed mutagenesis, we identified mutations that reduced mouse Bid lipid binding in vitro.Mutation of the conserved residue Lys157 specifically decreased the binding to negatively charged lipidsrelated to cardiolipin and additionally affected the rate of caspase cleavage. However, this lipid-bindingmutant had no discernable effect on Bid pro-apoptotic function in vivo. The results are interpreted in relationto an underlying interaction of Bid with lysophosphatidylcholine, which is not disrupted in any mutantretaining pro-apoptotic function both in vitro and in vivo.