Reversible membrane interaction of BAD requires two C-terminal lipid binding domains in conjunction with 14-3-3 protein binding
Reversible membrane interaction of BAD requires two C-terminal lipid binding domains in conjunction with 14-3-3 protein binding
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DOI：
10.1074/jbc.m600292200
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发表时间：
2006-06-23
影响因子：
4.8
通讯作者：
Rapp, Ulf R.
中科院分区：
文献类型：
--
作者：
Hekman, Mirko;Albert, Stefan;Rapp, Ulf R.
BAD是Bcl-2蛋白家族的仅Bcl-2同源结构域3（BH 3）的促凋亡成员，其响应于存活因子通过磷酸化调节。BAD与线粒体的结合被认为仅由其BH 3结构域介导。我们在这里表明，BAD结合脂质具有高亲和力，主要是带负电荷的磷脂，如磷脂酰丝氨酸，磷脂酸，心磷脂，以及富含胆固醇的脂质体。两个脂质结合结构域（LBD 1和LBD 2）具有不同的结合偏好被确定，都位于BAD蛋白的C-末端部分。BAD在需要LBD 2的过程中促进Bcl-X-L的膜易位。LBD 1和LBD 2的完整性也是体内促凋亡活性所必需的。BAD的磷酸化不影响膜结合，但使BAD对14-3-3蛋白的膜提取敏感。BAD可以通过14-3-3 zeta、-eta、-tau有效去除，而通过其他14-3-3同种型效率较低。组装的BAD中心点14-3-3复合物对富含胆固醇的脂质体表现出高亲和力，但对线粒体膜表现出低亲和力。我们的结论是，BAD是一种膜相关蛋白，具有受体而不是配体的标志。脂质结合对于BAD在体内的促凋亡功能是必不可少的。这些数据支持一种模型，其中BAD以磷酸化依赖的方式在线粒体和其他膜之间穿梭，并且14-3-3是这种重新定位的关键调节因子。BAD与膜的动态相互作用与Bcl-X-L的活化和膜移位有关。
BAD is a Bcl-2 homology domain 3 (BH3)-only proapoptotic member of the Bcl-2 protein family that is regulated by phosphorylation in response to survival factors. Binding of BAD to mitochondria is thought to be exclusively mediated by its BH3 domain. We show here that BAD binds to lipids with high affinities, predominantly to negatively charged phospholipids, such as phosphatidylserine, phosphatidic acid, and cardiolipin, as well as to cholesterol-rich liposomes. Two lipid binding domains (LBD1 and LBD2) with different binding preferences were identified, both located in the C-terminal part of the BAD protein. BAD facilitates membrane translocation of Bcl-X-L in a process that requires LBD2. Integrity of LBD1 and LBD2 is also required for proapoptotic activity in vivo. Phosphorylation of BAD does not affect membrane binding but renders BAD susceptible to membrane extraction by 14-3-3 proteins. BAD can be removed efficiently by 14-3-3 zeta, -eta, -tau and less efficiently by other 14-3-3 isoforms. The assembled BAD center dot 14-3-3 complex exhibited high affinity for cholesterol-rich liposomes but low affinity for mitochondrial membranes. We conclude that BAD is a membrane-associated protein that has the hallmarks of a receptor rather than a ligand. Lipid binding is essential for the proapoptotic function of BAD in vivo. The data support a model in which BAD shuttles in a phosphorylation-dependent manner between mitochondria and other membranes and where 14-3-3 is a key regulator of this relocation. The dynamic interaction of BAD with membranes is tied to activation and membrane translocation of Bcl-X-L.