Molecular analysis of functional redundancy among anti-apoptotic Bcl-2 proteins and its role in cancer cell survival.
Molecular analysis of functional redundancy among anti-apoptotic Bcl-2 proteins and its role in cancer cell survival.
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DOI：
10.1016/j.yexcr.2014.02.010
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发表时间：
2014-04-01
影响因子：
3.7
通讯作者：
Chambers, Timothy C.
中科院分区：
文献类型：
--
作者：
Eichhorn, Joshua M.;Alford, Sarah E.;Sakurikar, Nandini;Chambers, Timothy C.
Bcl-2家族蛋白是内源性细胞凋亡的重要调节因子和介质。一些证据表明，该家族的抗凋亡成员，包括Bcl-2，Bcl-xL和Mcl-1，表现出功能冗余。然而，目前的证据主要是间接的，主要基于使用小分子抑制剂的药理学数据。为了在分子水平上研究抗凋亡Bcl-2蛋白的补偿和冗余，我们使用了组合敲低/过表达策略，基本上用另一个成员取代一个成员的功能。结果表明，HeLa细胞严格依赖于Mcl-1的生存和相应的难治性Bcl-2/Bcl-xL抑制剂ABT-263，并保持耐ABT-263的Bcl-xL过表达的背景下，因为内源性Mcl-1继续提供主要的监护人的作用。然而，如果Mcl-1在Bcl-xL过表达的情况下被敲低，则细胞变得Bcl-xL依赖性并且对ABT-263敏感。我们还表明，Bcl-xL通过隔离两个关键的促凋亡Bcl-2家族成员，巴克和Bim（通常与Mcl-1结合）来补偿Mcl-1的损失，并且Bim对于Mcl-1敲低诱导的细胞死亡是必需的。据我们所知，这是第一个例子，其中Mcl-1的损失诱导的细胞死亡是由一个单一的BH 3-只有Bcl-2家族成员的沉默拯救。在结肠癌细胞系中，Bcl-xL和Mcl-1也发挥代偿作用，并且Mcl-1敲低使细胞对ABT-263敏感。采用敲除和过表达相结合的新策略获得的结果为促生存Bcl-2家族蛋白的补偿机制提供了独特的分子见解。
Bcl-2 family proteins act as essential regulators and mediators of intrinsic apoptosis. Several lines of evidence suggest that the anti-apoptotic members of the family, including Bcl-2, Bcl-xL and Mcl-1, exhibit functional redundancy. However, the current evidence is largely indirect, and based mainly on pharmacological data using small-molecule inhibitors. In order to study compensation and redundancy of anti-apoptotic Bcl-2 proteins at the molecular level, we used a combined knockdown/overexpression strategy to essentially replace the function of one member with another. The results show that HeLa cells are strictly dependent on Mcl-1 for survival and correspondingly refractory to the Bcl-2/Bcl-xL inhibitor ABT-263, and remain resistant to ABT-263 in the context of Bcl-xL overexpression because endogenous Mcl-1 continues to provide the primary guardian role. However, if Mcl-1 is knocked down in the context of Bcl-xL overexpression, the cells become Bcl-xL-dependent and sensitive to ABT-263. We also show that Bcl-xL compensates for loss of Mcl-1 by sequestration of two key pro-apoptotic Bcl-2 family members, Bak and Bim, normally bound to Mcl-1, and that Bim is essential for cell death induced by Mcl-1 knockdown. To our knowledge, this is the first example where cell death induced by loss of Mcl-1 was rescued by the silencing of a single BH3-only Bcl-2 family member. In colon carcinoma cell lines, Bcl-xL and Mcl-1 also play compensatory roles, and Mcl-1 knockdown sensitizes cells to ABT-263. The results, obtained employing a novel strategy of combining knockdown and overexpression, provide unique molecular insight into the mechanisms of compensation by pro-survival Bcl-2 family proteins.
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