Single-point mutations of a lysine residue change function of Bax and Bcl-xL expressed in Bax- and Bak-less mouse embryonic fibroblasts: novel insights into the molecular mechanisms of Bax- induced apoptosis
Single-point mutations of a lysine residue change function of Bax and Bcl-xL expressed in Bax- and Bak-less mouse embryonic fibroblasts: novel insights into the molecular mechanisms of Bax- induced apoptosis
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DOI：
10.1038/cdd.2010.112
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发表时间：
2011-03-01
影响因子：
12.4
通讯作者：
Gulbins, E.
中科院分区：
文献类型：
--
作者：
Szabo, I.;Soddemann, M.;Gulbins, E.
Bcl-2家族的成员在促凋亡中发挥着关键作用（例如，例如，在一个实施例中，Bax）和抗凋亡（e.例如，在一个实施例中，Bcl-x（L））调节程序性细胞死亡。我们以前确定线粒体钾通道Kv1.3作为Bax的新靶点。孵育Kv1.3阳性分离的线粒体与Bax引发凋亡事件，而Kv1.3缺陷的线粒体抵抗这种刺激。Bax在赖氨酸128位的突变（Bax K128 E）消除了其对Kv1.3的作用和对线粒体凋亡变化的诱导。这些数据表明Bax对Kv1.3的毒素样作用，以触发至少一些典型的细胞凋亡的线粒体变化。为了深入了解Bcl-Kv 1.3相互作用的机制，我们将Bcl-x（L）的Glu 158（对应于Bax中的K128）突变为赖氨酸。这种取代使Bcl-x（L）促凋亡。用野生型Bax、Bcl-x（L）E158 E或Bcl-x（L）E158 K转染双敲除（Bax（-/-）/巴克（-/-））小鼠胚胎成纤维细胞（DKO MEFs）显示，在DKO MEFs和Bcl-x E128 E转染的细胞中，由各种刺激诱导的凋亡是缺陷的，但在用Bcl-x E158 K或野生型Bax转染后恢复。野生型Bax和BaxK 128 E在掺入平面脂质双层后都可以形成类似的离子导电孔。我们的结果表明Bax与Kv1.3存在生理相关的相互作用，并进一步表明不同的赖氨酸在决定Bcl 2家族蛋白的促细胞凋亡特征方面发挥着至关重要的作用。Cell Death and Differentiation（2011）18，427-438; doi：10.1038/cdd.2010.112; 2010年10月1日在线发表
Members of the Bcl-2 family play key roles as proapoptotic (e. g., Bax) and antiapoptotic (e. g., Bcl-x(L)) regulators of programmed cell death. We previously identified the mitochondrial potassium channel Kv1.3 as a novel target of Bax. Incubating Kv1.3-positive isolated mitochondria with Bax triggered apoptotic events, whereas Kv1.3-deficient mitochondria were resistant to this stimulus. Mutation of Bax at lysine 128 (BaxK128E) abrogated its effects on Kv1.3 and the induction of apoptotic changes in mitochondria. These data indicate a toxin-like action of Bax on Kv1.3 to trigger at least some of the mitochondrial changes typical for apoptosis. To gain insight into the mechanism of Bax-Kv1.3 interaction, we mutated Glu158 of Bcl-x(L) (corresponding to K128 in Bax) to lysine. This substitution turned Bcl-x(L) proapoptotic. Transfection of double knockout (Bax(-/-)/Bak(-/-)) mouse embryonic fibroblasts (DKO MEFs) with either wild-type Bax, BaxK128E, or Bcl-x(L)E158K showed that apoptosis induced by various stimuli was defective in DKO MEFs and BaxK128E-transfected cells, but was recovered upon transfection with Bcl-xLE158K or wild-type Bax. Both wild-type Bax and BaxK128E can form similar ion-conducting pores upon incorporation into planar lipid bilayers. Our results point to a physiologically relevant interaction of Bax with Kv1.3 and further indicate a crucial role of a distinct lysine in determining the proapoptotic character of Bcl2-family proteins. Cell Death and Differentiation (2011) 18, 427-438; doi:10.1038/cdd.2010.112; published online 1 October 2010