Moreira Douglas de Souza, Duarte Ana Paula, Pais Fabiano Sviatopolk Mirsky, Silva-Pereira Rosiane Aparecida da, Romanha Alvaro José, Schenkman Sergio, Murta Silvane Maria Fonseca
Fundação Oswaldo Cruz-Fiocruz, Instituto René Rachou, Belo Horizonte, MG, Brasil.
Universidade Federal de São Paulo, Departamento de Microbiologia, Imunologia e Parasitologia, São Paulo, SP, Brasil.
Mem Inst Oswaldo Cruz. 2018 Jul 26;113(9):e180162. doi: 10.1590/0074-02760180162.
Eukaryotic initiation factor 5A (eIF5A) is a conserved protein with an essential role in translation elongation. Using one and two-dimensional western blotting, we showed that the eIF5A protein level was 2-fold lower in benznidazole (BZ)-resistant (BZR and 17LER) Trypanosoma cruzi populations than in their respective susceptible counterparts (BZS and 17WTS). To confirm the role of eIF5A in BZ resistance, we transfected BZS and 17WTS with the wild-type eIF5A or mutant eIF5A-S2A (in which serine 2 was replaced by alanine). Upon overexpressing eIF5A, both susceptible lines became approximately 3- and 5-fold more sensitive to BZ. In contrast, the eIF5A-S2A mutant did not alter its susceptibility to BZ. These data suggest that BZ resistance might arise from either decreasing the translation of proteins that require eIF5A, or as a consequence of differential levels of precursors for the hypusination reactions (e.g., spermidine and trypanothione), both of which alter BZ's effects in the parasite.
真核生物起始因子5A（eIF5A）是一种保守蛋白，在翻译延伸过程中发挥着重要作用。我们通过一维和二维蛋白质印迹法表明，在对苯硝唑（BZ）耐药的克氏锥虫群体（BZR和17LER）中，eIF5A蛋白水平比各自对应的敏感群体（BZS和17WTS）低2倍。为了证实eIF5A在BZ耐药中的作用，我们用野生型eIF5A或突变型eIF5A-S2A（其中丝氨酸2被丙氨酸取代）转染BZS和17WTS。过表达eIF5A后，两个敏感株系对BZ的敏感性分别提高了约3倍和5倍。相反，eIF5A-S2A突变体对BZ的敏感性没有改变。这些数据表明，BZ耐药可能源于需要eIF5A的蛋白质翻译减少，或者是由于hypusination反应前体（如亚精胺和锥虫硫醇）水平的差异，这两者都会改变BZ对寄生虫的作用。
