Cell Signalling and Cell Death Division andDepartment of Medical Biology, University of Melbourne, Parkville, VIC 3050, Australia
Cell Signalling and Cell Death Division andDepartment of Medical Biology, University of Melbourne, Parkville, VIC 3050, Australia.
Proc Natl Acad Sci U S A. 2014 Jun 10;111(23):8512-7. doi: 10.1073/pnas.1406425111. Epub 2014 May 27.
Antiapoptotic B-cell lymphoma 2 (Bcl-2) family members such as Bcl-2, myeloid cell leukemia 1 (Mcl-1), and B-cell lymphoma-X large (Bcl-xL) are proposed to inhibit autophagy by directly binding to the BH3 domain of Beclin 1/Atg6. However, these Bcl-2 family proteins also block the proapoptotic activity of Bcl-2-associated X (Bax) and Bcl-2 homologous antagonist/killer (Bak), and many inducers of autophagy also cause cell death. Therefore, when the mitochondrial-mediated apoptosis pathway is functional, interpretation of such experiments is complicated. To directly test the impact of the endogenous antiapoptotic Bcl-2 family members on autophagy in the absence of apoptosis, we inhibited their activity in cells lacking the essential cell death mediators Bax and Bak. We also used inducible lentiviral vectors to overexpress Bcl-2, Bcl-xL, or Mcl-1 in cells and subjected them to treatments that promote autophagy. In the absence of Bax and Bak, Bcl-2, Bcl-xL, and Mcl-1 had no detectable effect on autophagy or cell death in myeloid or fibroblast cell lines. On the other hand, when Bax and Bak were present, inhibiting the prosurvival Bcl-2 family members stimulated autophagy, but this correlated with increased cell death. In addition, inhibition of autophagy induced by amino acid starvation, etoposide, or interleukin-3 withdrawal did not affect cell death in the absence of Bax and Bak. These results demonstrate that the antiapoptotic Bcl-2 family members do not directly inhibit components of the autophagic pathway but instead affect autophagy indirectly, owing to their inhibition of Bax and Bak.
凋亡抑制蛋白 B 细胞淋巴瘤 2（Bcl-2）家族成员，如 Bcl-2、髓样细胞白血病 1（Mcl-1）和 B 细胞淋巴瘤-X 长（Bcl-xL），被认为通过直接与 Beclin 1/Atg6 的 BH3 结构域结合来抑制自噬。然而，这些 Bcl-2 家族蛋白也阻止了 Bcl-2 相关 X（Bax）和 Bcl-2 同源拮抗剂/杀伤（Bak）的促凋亡活性，并且许多自噬诱导剂也会导致细胞死亡。因此，当线粒体介导的凋亡途径功能正常时，这些实验的解释就变得复杂了。为了直接测试内源性抗凋亡 Bcl-2 家族成员在没有凋亡的情况下对自噬的影响，我们在缺乏必需的细胞死亡介质 Bax 和 Bak 的细胞中抑制了它们的活性。我们还使用可诱导的慢病毒载体在细胞中过表达 Bcl-2、Bcl-xL 或 Mcl-1，并对它们进行促进自噬的处理。在缺乏 Bax 和 Bak 的情况下，Bcl-2、Bcl-xL 和 Mcl-1 对髓样或成纤维细胞系中的自噬或细胞死亡没有可检测到的影响。另一方面，当 Bax 和 Bak 存在时，抑制生存促进的 Bcl-2 家族成员刺激自噬，但这与增加的细胞死亡相关。此外，氨基酸饥饿、依托泊苷或白细胞介素-3 缺失诱导的自噬抑制不会影响 Bax 和 Bak 缺失时的细胞死亡。这些结果表明，抗凋亡 Bcl-2 家族成员不会直接抑制自噬途径的成分，而是通过抑制 Bax 和 Bak 间接影响自噬。
