Center of Excellence for Molecular Biology and Genomics of Shrimp, Department of Biochemistry, Faculty of Science, Chulalongkorn University, Thailand.
Center of Excellence for Molecular Biology and Genomics of Shrimp, Department of Biochemistry, Faculty of Science, Chulalongkorn University, Thailand; Omics Science and Bioinformatics Center, Faculty of Science, Chulalongkorn University, Thailand.
Fish Shellfish Immunol. 2018 Oct;81:284-296. doi: 10.1016/j.fsi.2018.06.054. Epub 2018 Jul 18.
While toxin-harboring Vibrio parahaemolyticus has been previously established as the causative agent of early mortality syndrome (EMS) or acute hepatopancreatic necrosis disease (AHPND) in shrimp, information on the mechanistic processes that happen in the host during infection is still lacking. Here, we examined the expression responses of the shrimp hemocyte transcriptome to V. parahaemolyticus AHPND (VP) by RNA sequencing (RNA-seq). Using libraries (SRA accession number SRP137285) prepared from shrimp hemocytes under experimental conditions, a reference library was de novo assembled for gene expression analysis of VP-challenged samples at 0, 3/6, and 48 h post infection (hpi). Using the library from 0-hpi as the control, 359 transcripts were found to be differentially expressed in the 3/6-hpi library, while 429 were differentially expressed in the 48-hpi library. The expression patterns reported in the RNA-seq of 9 representative genes such as anti-lipopolysaccharide factor (LvALF), crustin p (CRU), serpin 3 (SER), C-type lectin 3 (CTL), clottable protein 2 (CLO), mitogen-activated protein kinase kinase 4 (MKK4), P38 mitogen-activated protein kinase (P38), protein kinase A regulatory subunit 1 (PKA) and DNAJ homolog subfamily C member 1-like (DNJ) were validated by qRT-PCR. The expression of these genes was also analyzed in shrimp that were injected with the partially purified VP toxin. A VP toxin-responsive gene, LvALF was identified, and its function was characterized by RNA interference. LvALF knockdown resulted in significantly rapid increase of shrimp mortality caused by toxin injection. Protein-protein interaction analysis by molecular docking suggested that LvALF possibly neutralizes VP toxin through its LPS-binding domain. The data generated in this study provide preliminary insights into the differences in the immune response of shrimp to the bacterial and toxic aspect of VP as a disease.
尽管产毒副溶血弧菌先前被确定为虾早期死亡综合征 (EMS) 或急性肝胰腺坏死病 (AHPND) 的病原体，但宿主在感染过程中发生的机制过程的信息仍然缺乏。在这里，我们通过 RNA 测序 (RNA-seq) 检查了虾血细胞转录组对副溶血弧菌 AHPND (VP) 的表达反应。使用在实验条件下从虾血细胞制备的文库 (SRA 访问号 SRP137285)，从头组装参考文库以用于分析 VP 感染样品在 0、3/6 和 48 小时攻毒后的基因表达。使用 0 小时攻毒的文库作为对照，在 3/6 小时攻毒文库中发现 359 个转录物差异表达，而在 48 小时攻毒文库中发现 429 个转录物差异表达。RNA-seq 中报告的 9 个代表性基因（如抗脂多糖因子 (LvALF)、甲壳质 p (CRU)、丝氨酸蛋白酶抑制剂 3 (SER)、C 型凝集素 3 (CTL)、凝结蛋白 2 (CLO)、丝裂原活化蛋白激酶激酶 4 (MKK4)、P38 丝裂原活化蛋白激酶 (P38)、蛋白激酶 A 调节亚基 1 (PKA) 和 DNAJ 同源亚家族 C 成员 1 样 (DNJ) 的表达模式通过 qRT-PCR 进行了验证。还分析了用部分纯化的 VP 毒素注射的虾中这些基因的表达。鉴定了一个 VP 毒素反应基因 LvALF，并通过 RNA 干扰对其功能进行了表征。LvALF 敲低导致毒素注射引起的虾死亡率显著快速增加。分子对接的蛋白质-蛋白质相互作用分析表明，LvALF 可能通过其 LPS 结合结构域中和 VP 毒素。本研究产生的数据初步揭示了虾对 VP 细菌和毒性方面的免疫反应差异作为一种疾病。
