Norwegian College of Fishery Science, Faculty of Biosciences, Fisheries & Economics, UiT The Arctic University of Norway, Tromsø, Norway.
Department of Physics and Technology, UiT The Arctic University of Norway, Tromsø, Norway.
Fish Shellfish Immunol. 2020 Apr;99:119-129. doi: 10.1016/j.fsi.2020.01.058. Epub 2020 Jan 31.
Rab GTPases control trafficking of intracellular vesicles and are key regulators of endocytic and secretory pathways. Due to their specific distribution, they may serve as markers for different endolysosomal compartments. Since Rab GTPases are involved in uptake and trafficking of endocytosed ligands and cell receptors, as well as secretion of immune mediators, they have been implicated in diverse immunological processes and their functions are often exploited by intracellular pathogens such as viruses. While Rab proteins have been studied extensively in mammals, their functions in vesicle trafficking in teleosts are not well known. In the present work, Atlantic salmon Rab5c, Rab7a and Rab27a homologs were studied in terms of intracellular distribution and gene expression. Structured illumination microscopy demonstrated that transgenic, GFP-tagged salmon Rab5c and Rab7a are, predominantly, located within early endosomes and late endosomes/lysosomes, respectively. In contrast, Rab27a showed a broader distribution, which indicates that it associates with diverse intracellular vesicles and organelles. Infection of salmon with Salmonid alphavirus subtype 3 (SAV3) enhanced the mRNA levels of all of the studied Rab isoforms in heart and head kidney and most of them were upregulated in spleen. This may reflect the capacity of the virus to exploit the functions of these rab proteins. It is also possible that the transcriptional regulation of Rab proteins in SAV3-infected organs may play a role in the antiviral immune response. The latter was further supported by in vitro experiments with adherent head kidney leukocytes. The expression of Rab5c and Rab27a was upregulated in these cells following stimulation with TLR ligands including CpG oligonucleotides and polyI:C. The expression of most of the analyzed Rab isoforms in the primary leukocytes was also enhanced by stimulation with type I IFN. Interestingly, IFN-gamma had a negative effect on Rab7a expression which may be linked to the priming activity of this cytokine on monocytes and macrophages. Overall, these data demonstrate that the intracellular distribution of Rab5c, Rab7a and Rab27a is phylogenetically conserved within vertebrates and that these molecules might be implicated in viral infections and the regulation of the antiviral immune response in Atlantic salmon.
Rab GTPases 控制细胞内囊泡的运输，是内吞和分泌途径的关键调节因子。由于它们的特定分布，它们可以作为不同内溶酶体隔室的标志物。由于 Rab GTPases 参与内吞配体和细胞受体的摄取和运输以及免疫介质的分泌，它们与多种免疫过程有关，其功能经常被细胞内病原体如病毒利用。虽然 Rab 蛋白在哺乳动物中得到了广泛的研究，但它们在硬骨鱼中的囊泡运输功能尚不清楚。在本工作中，研究了大西洋鲑 Rab5c、Rab7a 和 Rab27a 同源物的细胞内分布和基因表达。结构照明显微镜表明，转染的 GFP 标记的鲑 Rab5c 和 Rab7a 主要位于早期内体和晚期内体/溶酶体中。相比之下，Rab27a 表现出更广泛的分布，这表明它与不同的细胞内囊泡和细胞器相关。鲑鱼感染鲑鱼甲病毒 3 亚型（SAV3）后，所有研究的 Rab 同工型的 mRNA 水平在心和头肾中均增强，其中大多数在脾中上调。这可能反映了病毒利用这些 rab 蛋白功能的能力。也可能是 SAV3 感染器官中 Rab 蛋白的转录调节在抗病毒免疫反应中发挥作用。后者进一步得到了与粘附性头肾白细胞的体外实验的支持。这些细胞中 Rab5c 和 Rab27a 的表达在 TLR 配体（包括 CpG 寡核苷酸和聚 I：C）刺激后上调。大多数分析的 Rab 同工型在原代白细胞中的表达也被 I 型 IFN 刺激增强。有趣的是，IFN-γ对 Rab7a 表达有负作用，这可能与该细胞因子对单核细胞和巨噬细胞的启动活性有关。总的来说，这些数据表明 Rab5c、Rab7a 和 Rab27a 的细胞内分布在脊椎动物中是进化保守的，这些分子可能与大西洋鲑的病毒感染和抗病毒免疫反应的调节有关。
