Yarbrough Azure, Maringer Katherine, Saheb Entsar J, Jawed Sanaa, Bush John
Biology Department, University of Arkansas at Little Rock, 2801 South University Ave, Little Rock, AR 72204-1099, USA.
Otsuka Pharmaceuticals, Greater Pittsburg Area, Pittsburgh, PA 15201, USA.
Biology (Basel). 2018 May 28;7(2):33. doi: 10.3390/biology7020033.
Rab GTPases are essential regulators of many cellular processes and play an important role in downstream signaling vital to proper cell function. We sought to elucidate the role of novel GTPase RabS. Cell lines over-expressing DdRabS and expressing DdRabS N137I (dominant negative (DN)) proteins were generated, and it was determined that DdRabS localized to endosomes, ER-Golgi membranes, and the contractile vacuole system. It appeared to function in vesicular trafficking, and the secretion of lysosomal enzymes. Interestingly, microscopic analysis of GFP-tagged DdRabS (DN) cells showed differential localization to lysosomes and endosomes compared to GFP-tagged DdRabS overexpressing cells. Both cell lines over-secreted lysosomal glycosidase enzymes, especially β-glucosidase. Furthermore, DdRabS overexpressing cells were defective in aggregation due to decreased cell⁻cell cohesion and sensitivity to cAMP, leading to abnormal chemotactic migration, the inability to complete development, and increased induced cell death. These data support a role for DdRabS in trafficking along the vesicular and biosynthetic pathways. We hypothesize that overexpression of DdRabS may interfere with GTP activation of related proteins essential for normal development resulting in a cascade of defects throughout these processes.
Rab GTP 酶是许多细胞过程的重要调节因子，在对细胞正常功能至关重要的下游信号传导中发挥重要作用。我们试图阐明新型 GTP 酶 RabS 的作用。构建了过表达 DdRabS 和表达 DdRabS N137I（显性负性（DN））蛋白的细胞系，确定 DdRabS 定位于内体、内质网 - 高尔基体膜和收缩泡系统。它似乎在囊泡运输和溶酶体酶的分泌中发挥作用。有趣的是，与过表达 GFP 标记的 DdRabS 的细胞相比，对 GFP 标记的 DdRabS（DN）细胞的显微镜分析显示其在溶酶体和内体中的定位不同。两种细胞系都过度分泌溶酶体糖苷酶，尤其是β - 葡萄糖苷酶。此外，过表达 DdRabS 的细胞由于细胞间黏附力降低和对 cAMP 的敏感性而在聚集方面存在缺陷，导致异常的趋化性迁移、无法完成发育以及诱导细胞死亡增加。这些数据支持 DdRabS 在沿囊泡和生物合成途径的运输中发挥作用。我们假设 DdRabS 的过表达可能会干扰正常发育所必需的相关蛋白的 GTP 激活，从而在这些过程中导致一系列缺陷。
