DataSheet1_The integrated stress response protects against ER stress but is not required for altered translation and lifespan from dietary restriction in Caenorhabditis elegans.docx

The highly conserved integrated stress response (ISR) reduces and redirects mRNA translation in response to certain forms of stress and nutrient limitation. It is activated when kinases phosphorylate a key residue in the alpha subunit of eukaryotic translation initiation factor 2 (eIF2). General Control Nonderepressible-2 (GCN2) is activated to phosphorylate eIF2α by the presence of uncharged tRNA associated with nutrient scarcity, while protein kinase R-like ER kinase-1 (PERK) is activated during the ER unfolded protein response (UPRER). Here, we investigated the role of the ISR during nutrient limitation and ER stress with respect to changes in protein synthesis, translationally driven mRNA turnover, and survival in Caenorhabditis elegans. We found that, while GCN2 phosphorylates eIF2α when nutrients are restricted, the ability to phosphorylate eIF2α is not required for changes in translation, nonsense-mediated decay, or lifespan associated with dietary restriction (DR). Interestingly, loss of both GCN2 and PERK abolishes increased lifespan associated with dietary restriction, indicating the possibility of other substrates for these kinases. The ISR was not dispensable under ER stress conditions, as demonstrated by the requirement for PERK and eIF2α phosphorylation for decreased translation and wild type-like survival. Taken together, results indicate that the ISR is critical for ER stress and that other translation regulatory mechanisms are sufficient for increased lifespan under dietary restriction.

高度保守的整合应激反应（integrated stress response, ISR）可在机体遭遇特定应激与营养限制时，下调并重定向mRNA翻译过程。当真核翻译起始因子2（eukaryotic translation initiation factor 2, eIF2）α亚基的关键残基被激酶磷酸化时，该通路即被激活。通用控制非阻遏2（General Control Nonderepressible-2, GCN2）可通过感知与营养匮乏相关的未氨酰化tRNA，介导eIF2α的磷酸化；而蛋白激酶R样内质网激酶1（protein kinase R-like ER kinase-1, PERK）则在内质网未折叠蛋白反应（ER unfolded protein response, UPRER）过程中被激活。本研究以秀丽隐杆线虫（Caenorhabditis elegans）为模型，探究了整合应激反应在营养限制与内质网应激条件下，对蛋白质合成、翻译依赖性mRNA降解以及个体存活的调控作用。研究发现，尽管营养受限时GCN2可磷酸化eIF2α，但饮食限制（dietary restriction, DR）相关的翻译模式改变、无义介导的mRNA降解（nonsense-mediated decay, NMD）以及寿命延长，均不依赖于eIF2α的磷酸化过程。值得注意的是，同时缺失GCN2与PERK会完全阻断饮食限制介导的寿命延长，提示这两种激酶可能存在其他下游底物。在内质网应激条件下，整合应激反应并非可有可无：降低翻译水平与维持野生型个体存活率，均依赖于PERK和eIF2α的磷酸化，这一实验结果证实了该结论。综上，本研究结果表明，整合应激反应对内质网应激至关重要，而其他翻译调控机制足以介导饮食限制下的寿命延长。