Entamoeba histolytica is an intestinal parasite infecting over 50 million people worldwide and is the causative agent of amebic dysentery and amoebic liver abscess. In the human host, E. histolytica experiences stress brought on by nutrient deprivation and the host immune response. To be a successful parasite, E. histolytica must counter the stress; therefore, understanding the stress response may uncover new drug targets. In many systems, the stress response includes down-regulation of protein translation, which is regulated by phosphorylation of eukaryotic initiation factor (eIF-2α). Previous work has demonstrated that phosphorylation of the E. histolytica eIF-2α (EheIF-2α) increases significantly when exposed to long-term serum starvation, oxidative stress, and long-term heat shock. However, the effects of reagents that are known to induce nitrosative or endoplasmic reticulum (ER) stresses, on EheIF-2α have yet to be evaluated. Nitrosative stress is part of the host’s immune response and ER stress can be caused by several physiological or pathological factors. We treated E. histolytica cells with various reagents known to induce nitrosative stress (DPTA-NONOate and SNP) or ER stress (BFA and DTT). We examined the morphology of the ER, tracked phosphorylation of EheIF-2α, and assessed protein translation in control and stressed cells. While all four stress-inducing reagents caused a global reduction in protein translation, only DTT was capable of also inducing changes in the morphology of the ER (consistent with ER stress) and phosphorylation of EheIF-2α. This suggests that DTT authentically induces ER stress in E. histolytica and that this stress is managed by the eIF-2α-based system. This was supported by the observation that cells expressing a non-phosphorylatable version of eIF-2α were also highly sensitive to DTT-stress. Since protein translation decreased in the absence of phosphorylation of eIF-2α (after treatment with DPTA-NONOate, SNP or BFA), the data also indicate that there are alternative protein-translational control pathways in E. histolytica. Overall, our study further illuminates the stress response to nitrosative stress and ER stress in E. histolytica.

溶组织内阿米巴是一种肠道寄生虫，感染全世界超过 5000 万人，是阿米巴痢疾和阿米巴肝脓肿的病原体。在人类宿主中，溶组织内阿米巴会经历营养缺乏和宿主免疫反应带来的压力。为了成为成功的寄生虫，溶组织内阿米巴必须对抗压力。因此，了解应激反应可能会发现新的药物靶点。在许多系统中，应激反应包括蛋白质翻译的下调，这是由真核起始因子 (eIF-2α) 的磷酸化调节的。先前的工作表明，当暴露于长期血清饥饿、氧化应激和长期热休克时，溶组织内阿米巴eIF-2α ( Eh eIF-2α) 的磷酸化显着增加。然而，已知可诱导亚硝化或内质网 (ER) 应激的试剂对Eh eIF-2α 的影响尚未得到评估。亚硝化应激是宿主免疫反应的一部分，内质网应激可由多种生理或病理因素引起。我们用已知可诱导亚硝化应激（DPTA-NONOate 和 SNP）或 ER 应激（BFA 和 DTT）的各种试剂处理溶组织内阿米巴细胞。我们检查了 ER 的形态，跟踪了Eh eIF-2α 的磷酸化，并评估了对照细胞和应激细胞中的蛋白质翻译。虽然所有四种应激诱导试剂都会导致蛋白质翻译的整体减少，但只有 DTT 还能够诱导 ER 形态的变化（与 ER 应激一致）和Eh eIF-2α 的磷酸化。这表明 DTT 确实诱导了大肠杆菌的 ER 应激。 histolytica 并且这种压力是由基于 eIF-2α 的系统管理的。表达非磷酸化版本 eIF-2α 的细胞对 DTT 应激也高度敏感，这一观察结果支持了这一点。由于在没有 eIF-2α 磷酸化的情况下（用 DPTA-NONOate、SNP 或 BFA 处理后）蛋白质翻译减少，因此数据还表明，溶组织内阿米巴中存在替代的蛋白质翻译控制途径。总体而言，我们的研究进一步阐明了溶组织内阿米巴对亚硝化应激和内质网应激的应激反应。