小龙虾prx6基因在对抗金黄色葡萄球菌感染中的分子作用机制研究  

Research on the Molecular Mechanism of Crayfish prx 6 in the Process of Defending against Staphylococcus aureus Infection

作　　者：金博阳 秦仕宇 张明达 李倩倩 文静 沈秀丽[2] 杜志强 JIN Bo-yang;QIN Shi-yu;ZHANG Ming-da;LI Qian-qian;WEN Jing;SHEN Xiu-i;DU Zhi-qiang(School of Life Science and Technology,Inner Mongolia University of Science and Technology,Baotou 014010;Inner Mongolia University of Science and Technology Library,Baotou 014010)

机构地区：[1]内蒙古科技大学生命科学与技术学院,包头014010 [2]内蒙古科技大学图书馆,包头014010

出　　处：《生物技术通报》2024年第7期314-322,共9页Biotechnology Bulletin

基　　金：内蒙古自治区自然科学基金项目(2024MS03051);国家自然科学基金项目(32060834);内蒙古科技大学2022年基本科研业务费专项资金(028)。

摘　　要：【目的】旨在研究prx 6基因在小龙虾先天免疫中的调控作用机制,探索小龙虾prx 6基因在金黄色葡萄球菌(Staphylococcus aureus)感染后发挥免疫调控作用的可能分子机制。【方法】选取prx 6基因作为研究对象,提取正常小龙虾各组织的总RNA,利用RT-qPCR分析目的基因在小龙虾各组织中的分布,并检测S.aureus免疫刺激后,prx 6基因在小龙虾相关组织中的表达模式。通过RNA干扰技术(RNAi)敲低prx 6基因表达量,在感染S.aureus后,测定小龙虾肝胰腺中Pc-crustin 3、Pc-crustin 4、Pc-ALF 9和Pc-lectin 1免疫效应基因的相对表达量。同时,进一步对小龙虾血淋巴中的细菌载量进行统计分析。【结果】通过对prx 6基因在小龙虾各组织的表达水平分析,观察到该基因在肝胰腺的表达量显著高于其他组织。在S.aureus感染后,小龙虾肝胰腺、血细胞和肠组织中prx 6基因的表达量显著上调,在鳃组织仅有早期表达量有所增加。小龙虾存活率的检测结果表明RNAi下调prx 6基因表达量之后,其生存率显著低于dsGFP+S.aureus对照组。为了进一步探索死亡率升高的原因,研究了抗菌肽基因在抗细菌防御中的表达水平。RNAi实验结果显示,在prx 6基因表达量降低的情况下,小龙虾肝胰腺Pc-crustin 3、Pc-crustin 4、Pc-ALF 9以及Pc-lectin 1基因表达水平都出现了显著下降。此外,小龙虾血淋巴中的细菌载量检测结果显示,prx 6基因RNAi组的细菌载量显著高于dsGFP+S.aureus对照组。【结论】小龙虾prx 6基因通过影响抗菌肽基因的表达,以及血淋巴细菌清除能力来参与抗细菌先天免疫应答。【Objective】To unravel the molecular intricacies associated with the immune modulatory role of the crayfish prx 6 gene in response to Staphylococcus aureus infection and to expound its function in the orchestration of innate immunity in crayfish,this investigation was carried out.【Method】The study pivoted on the prx 6 gene and harvested total RNA from diverse unstimulated crayfish tissues.The RT-qPCR was utilized to pinpoint the tissue-specific distribution patterns of this gene.Following immune provocation by S.aureus,the prx 6 gene expression dynamics within crayfish’s immune-centric tissues were monitored.The research further endeavored to gauge the relative expression of immune executor genes,specifically,Pc-crustin 3,Pc-crustin 4,Pc-ALF 9,and Pc-lectin 1 in the hepatopancreas after RNAi-mediated silencing of prx 6,in the wake of S.aureus infection.Additionally,a quantitative appraisal of the bacterial burden in crayfish hemolymph was undertaken.【Result】The investigations underscored an amplified expression of the prx 6 gene within the hepatopancreas as compared to other tissues.Post S.aureus infection,the prx 6 gene expression showcased substantial augmentation in the hepatopancreas,hemocytes,and intestinal tissues,with a preliminary surge of expression detected in gill tissues.In terms of survival rates,a pronounced decrement in crayfish vitality was observed after prx 6 knockdown,relative to dsGFP+S.aureus controls.To elucidate the key factors linked to escalated mortality,the expression strata of antimicrobial peptide genes integral to bacterial defense were quantified.The outcomes revealed a significant downturn in the expression of Pc-crustin 3,Pc-crustin 4,Pc-ALF 9,and Pc-lectin 1 within the hepatopancreas,seemingly tied to prx 6 repression.Intriguingly,assessment of hemolymph bacterial loads unveiled a substantial elevation in bacterial load in the prx 6 knockdown group as set against controls.【Conclusion】The prx 6 gene in crayfish plays a pivotal role in the antibacterial innate immu

关 键 词：先天免疫 过氧化物还原酶6 RNA干扰 抗菌肽 金黄色葡萄球菌 

分 类 号：S945.4[农业科学—水产养殖]