Department of Biomedical Chemistry, Graduate School of Medicine, The University of Tokyo, Japan.
PLoS Pathog. 2021 Feb 16;17(2):e1008909. doi: 10.1371/journal.ppat.1008909. eCollection 2021 Feb.
The eukaryotic translation initiation factor 5A (eIF5A) is a highly conserved protein and is essential in all eukaryotes. However, the specific roles of eIF5A in translation and in other biological processes remain elusive. In the present study, we described the role of eIF5A, its posttranslational modifications (PTM), and the biosynthetic pathway needed for the PTM in Entamoeba histolytica, the protozoan parasite responsible for amoebic dysentery and liver abscess in humans. E. histolytica encodes two isotypes of eIF5A and two isotypes of enzymes, deoxyhypusine synthase (DHS), responsible for their PTM. Both of the two eIF5A isotypes are functional, whereas only one DHS (EhDHS1, but not EhDHS2), is catalytically active. The DHS activity increased ~2000-fold when EhDHS1 was co-expressed with EhDHS2 in Escherichia coli, suggesting that the formation of a heteromeric complex is needed for full enzymatic activity. Both EhDHS1 and 2 genes were required for in vitro growth of E. histolytica trophozoites, indicated by small antisense RNA-mediated gene silencing. In trophozoites, only eIF5A2, but not eIF5A1, gene was actively transcribed. Gene silencing of eIF5A2 caused compensatory induction of expression of eIF5A1 gene, suggesting interchangeable role of the two eIF5A isotypes and also reinforcing the importance of eIF5As for parasite proliferation and survival. Furthermore, using a sibling species, Entamoeba invadens, we found that eIF5A1 gene was upregulated during excystation, while eIF5A2 was downregulated, suggesting that eIF5A1 gene plays an important role during differentiation. Taken together, these results have underscored the essentiality of eIF5A and DHS, for proliferation and potentially in the differentiation of this parasite, and suggest that the hypusination associated pathway represents a novel rational target for drug development against amebiasis.
真核翻译起始因子 5A（eIF5A）是一种高度保守的蛋白质，存在于所有真核生物中，是必不可少的。然而，eIF5A 在翻译和其他生物过程中的具体作用仍然难以捉摸。在本研究中，我们描述了真核生物翻译起始因子 5A（eIF5A）及其翻译后修饰（PTM）、以及该 PTM 生物合成途径在引起人类阿米巴痢疾和肝脓肿的原生动物寄生虫溶组织内阿米巴中的作用。溶组织内阿米巴编码两种 eIF5A 同工型和两种脱羟鸟氨酸合成酶（DHS）同工型，负责它们的 PTM。两种 eIF5A 同工型都是有功能的，而只有一种 DHS（EhDHS1，但不是 EhDHS2）具有催化活性。当 EhDHS1 与 EhDHS2 在大肠杆菌中共表达时，EhDHS 活性增加了约 2000 倍，这表明形成异源二聚体复合物是充分发挥酶活性所必需的。通过小干扰 RNA 介导的基因沉默，EhDHS1 和 2 基因都对溶组织内阿米巴滋养体的体外生长是必需的。在滋养体中，只有 eIF5A2，而不是 eIF5A1，基因被主动转录。eIF5A2 基因沉默导致 eIF5A1 基因的补偿性诱导表达，这表明两种 eIF5A 同工型可以互换，也进一步强调了 eIF5A 对寄生虫增殖和存活的重要性。此外，利用亲缘种 Entamoeba invadens，我们发现 eIF5A1 基因在脱囊过程中上调，而 eIF5A2 基因下调，这表明 eIF5A1 基因在分化过程中发挥重要作用。综上所述，这些结果强调了 eIF5A 和 DHS 在增殖和潜在分化中的重要性，提示与 hypusination 相关的途径可能是抗阿米巴病药物开发的一个新的合理靶点。
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