Functional study of kuruma shrimp immune-related genes

在这项研究中，我们利用dsRNA干扰技术阐明并鉴定了这些基因在体内的功能。转谷氨酰胺酶(Tgase)和凝血蛋白(CP)是参与对虾凝血级联反应的关键分子。我们利用双链RNA干扰技术(DsRNAi)在体内证明了转谷氨酰胺酶和凝血蛋白在对虾凝血系统中的功能参与，以及它们对抗菌肽和酚氧化酶原系统相关基因mRNA表达的影响。分别注射1μg和10μg的Tgase-g和CP-dsRNA后第1天，Tgase mRNA在鳃、心脏、血细胞、肝胰腺、肠和淋巴器官中的表达受到抑制，而CP基因仅在鳃和心脏组织中表达受到抑制。然而，在注射后第7天，从基因表达、凝血和蛋白质数据可以看出，基因和剂量都发生了系统性的基因沉默。白斑病毒和VibR…的挑战试验更多的带刺线虫揭示了Tgase和CP在对虾免疫系统中的关键作用。在Tgase和CP沉默后，抑制抗菌肽(壳氨酸和溶菌酶)和参与酚氧化酶原系统的基因(前酚氧化酶和〓-2巨球蛋白)，表明对虾的凝血和体液生物防御之间存在关联。这一发现表明，在螯合类和甲壳类动物中，抗菌肽和涉及酚氧化酶原系统的基因的激活可能存在另一种机制。综上所述，本研究1)为凝血和抗菌肽以及参与酚氧化酶原系统的基因之间的联系提供了证据，从而为这些分子的激活提供了另一种机制2)基因沉默效力的结果将为dsRNA干扰在对虾疾病和病原体的控制和管理中的应用提供一个预防措施。这项研究有助于更好地了解对虾免疫系统的基本生物学。较少

In this study, we elucidated and characterized the function of these genes in vivo using dsRNA interference. Transglutaminase (Tgase) and clotting protein (CP) are critical molecules involved in the blood coagulation cascade of shrimp. We demonstrate in vivo the functional involvement of transglutaminase and clotting protein in the shrimp blood coagulation system using double stranded RNA interference (dsRNAi) and their influence in the mRNA expression of antimicrobial peptides and genes involved in the prophenoloxidase system. Expression of Tgase mRNA was inhibited in gills, heart, hemocyte, hepatopancreas, intestine and lymphoid organ while the CP gene was suppressed only in gills and heart tissues on day-1 post injection with 1 μg and 10 μg of Tgase- and CP-dsRNA, respectively. However, at day-7 post injection, systemic gene silencing was observed for both genes and dosages as shown by mRNA expression, blood coagulation and protein data. Challenge test with white spot virus and Vibr … More io penaecida revealed the critical function of Tgase and CP in the immune system of shrimp. Suppression of antimicrobial peptides (crustine and lysozyme) and genes involved in the prophenoloxidase system (prophenoloxidase and 〓-2 macroglobulin) following Tgase and CP silencing demonstrate an association between blood coagulation and humoral biodefenses in shrimp. This finding suggests a possible alternative mechanism on the activation of antimicrobial peptides and genes involved in the prophenoloxidase system in chelicerates and crustaceans. In summary, this study 1)provided evidence on the association of blood coagulation and anti-microbial peptides and genes involved in the prophenoloxidase system and thus, presents an alternative mechanism on the activation of these molecules 2) gene silencing potency results will serve as a precaution on the application of dsRNA interference in the control and management of shrimp diseases and pathogens. This study contributed to the better understanding of the basic biology of the shrimp immune system. Less