The kinase PERK and the transcription factor ATF4 play distinct and essential roles in autophagy resulting from tunicamycin-induced ER stress
The kinase PERK and the transcription factor ATF4 play distinct and essential roles in autophagy resulting from tunicamycin-induced ER stress
复制标题
DOI：
10.1074/jbc.ra118.002829
复制
发表时间：
2019-05-17
影响因子：
4.8
通讯作者：
Engedal, Nikolai
中科院分区：
文献类型：
--
作者：
Luhr, Morten;Torgersen, Maria Lyngaas;Engedal, Nikolai
内质网（ER）应激被认为通过未折叠蛋白反应（UPR）介导的自噬机制组分的转录上调和微管相关蛋白1轻链3（LC 3）的调节来激活自噬。据报道，上游UPR组分胰腺EIF 2-激酶（PERK）和肌醇需要酶1（IRE 1）介导这些作用，表明UPR可能通过PERK和IRE 1刺激自噬。然而，普遍定期审议及其组成部分如何影响自噬活性尚未得到彻底研究。通过分析LC 3通过自噬途径的通量，以及自噬货物的隔离和降解，我们在这里最终表明，经典的ER应激子衣霉素（TM）增强哺乳动物细胞中的自噬活性。PERK及其下游因子，转录激活因子4（ATF 4），是这种诱导的关键，但令人惊讶的是，IRE 1组成性抑制自噬活性。TM诱导的自噬需要自噬相关蛋白13（ATG 13）、Unc-51样自噬激活激酶1/2（ULK 1/ULK 2）和GABA A型受体相关蛋白（GABARAPs），但有趣的是，LC 3蛋白似乎是多余的。引人注目的是，在LNCaP和HeLa细胞中，ATF 4独立于PERK被激活，我们进一步的研究表明，ATF 4和PERK通过不同的机制调节自噬。具体而言，ATF 4控制转录并且是自噬体形成所必需的，而PERK以不依赖于转录的方式起作用，并且在自噬途径中的螯合后步骤中是必需的。总之，我们的研究结果表明，TM诱导的UPR激活功能性自噬，而IRE 1是一个负调节器，PERK和ATF 4需要在自噬途径的不同步骤。
Endoplasmic reticulum (ER) stress is thought to activate autophagy via unfolded protein response (UPR)-mediated transcriptional up-regulation of autophagy machinery components and modulation of microtubule-associated protein 1 light chain 3 (LC3). The upstream UPR constituents pancreatic EIF2- kinase (PERK) and inositol-requiring enzyme 1 (IRE1) have been reported to mediate these effects, suggesting that UPR may stimulate autophagy via PERK and IRE1. However, how the UPR and its components affect autophagic activity has not been thoroughly examined. By analyzing the flux of LC3 through the autophagic pathway, as well as the sequestration and degradation of autophagic cargo, we here conclusively show that the classical ER stressor tunicamycin (TM) enhances autophagic activity in mammalian cells. PERK and its downstream factor, activating transcription factor 4 (ATF4), were crucial for this induction, but surprisingly, IRE1 constitutively suppressed autophagic activity. TM-induced autophagy required autophagy-related 13 (ATG13), Unc-51-like autophagy-activating kinases 1/2 (ULK1/ULK2), and GABA type A receptor-associated proteins (GABARAPs), but interestingly, LC3 proteins appeared to be redundant. Strikingly, ATF4 was activated independently of PERK in both LNCaP and HeLa cells, and our further examination revealed that ATF4 and PERK regulated autophagy through separate mechanisms. Specifically, whereas ATF4 controlled transcription and was essential for autophagosome formation, PERK acted in a transcription-independent manner and was required at a post-sequestration step in the autophagic pathway. In conclusion, our results indicate that TM-induced UPR activates functional autophagy, and whereas IRE1 is a negative regulator, PERK and ATF4 are required at distinct steps in the autophagic pathway.