Ubiquitin-like Atg8 protein is expressed during autophagy and the encystation process in Naegleria gruberi
Ubiquitin-like Atg8 protein is expressed during autophagy and the encystation process in Naegleria gruberi
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DOI：
10.1007/s00436-016-5293-x
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发表时间：
2017-01-01
影响因子：
2
通讯作者：
Shibayama, Mineko
中科院分区：
文献类型：
--
作者：
Cardenas-Zuniga, Roberto;Sanchez-Monroy, Virginia;Shibayama, Mineko
耐格里变形虫属的成员是自由生活的变形虫，迄今为止唯一描述的人类致病物种是福氏耐格里变形虫。然而，由于该物种的完整基因组尚未报道，因此采用非致病性格鲁伯猪笼草来描述福氏猪笼草的分子途径。无论如何，格鲁伯氏猪笼草中的某些机制（例如自噬）尚未得到表征。自噬参与一些原生动物的不同细胞过程，包括不必要的细胞器的回收、发育和细胞分化。在这项工作中，我们使用特定诱导剂雷帕霉素来表征格鲁伯氏球菌的自噬。通过超微结构分析观察到处理后的滋养体中自噬空泡的形成，实时定量PCR证明了atg8基因的过度表达。此外，我们使用 LysoTracker 检测到经处理的变形虫的液泡酸化增加。最后，利用共聚焦显微镜来识别雷帕霉素诱导的格鲁伯氏球菌滋养体细胞质中的Atg8蛋白信号，甚至在诱导包囊的滋养体中的Atg8蛋白信号。总之，N. gruberi 拥有 Atg8 蛋白同源物，该蛋白在雷帕霉素诱导的自噬机制以及这种自由生活的变形虫的包囊过程中过度表达。
Members of the Naegleria genus are free-living amoebae, and the only pathogenic specie described to date for humans is N. fowleri. However, as the complete genome of this specie has not been reported, nonpathogenic N. gruberi is employed to describe molecular pathways in N. fowleri. Regardless, certain mechanisms, such as autophagy, have not yet been characterized in N. gruberi. Autophagy is involved in different cellular processes in some protozoa, including the recycling of unnecessary organelles, development, and cell differentiation. In this work, we characterized autophagy in N. gruberi using the specific inducer rapamycin. The formation of autophagy vacuoles in treated trophozoites was observed by ultrastructural analysis, and real time quantitative PCR demonstrated overexpression of the atg8 gene. In addition, we detected an increase in the vacuolar acidification of treated amoebae using the LysoTracker. Finally, confocal microscopy was utilized to identify Atg8 protein signal in the cytoplasm of N. gruberi trophozoites induced with rapamycin and even in trophozoites induced to encyst. In conclusion, N. gruberi possesses an Atg8 protein homolog that is overexpressed during the autophagic mechanism induced by rapamycin and also during encystation of this free-living amoeba.