利用CRISPR/Cas9介导的斑马鱼基因敲除模型鉴定irf3抗病毒性出血性败血症病毒感染的功能特征

IF 4.6 Q2 MATERIALS SCIENCE, BIOMATERIALS
Sarithaa Raguvaran Sellaththurai , Sumi Jung , Kishanthini Nadarajapillai , Myoung-Jin Kim , Jehee Lee
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引用次数: 0

摘要

干扰素调节因子(IRFs)是一种转录因子,参与病原体反应调节、免疫细胞生长和分化等免疫反应。当病原体识别受体识别病毒 DNA 或 RNA 时,IRF 通过信号级联合成 I 型干扰素。我们发现,irf3 在早期胚胎阶段和成年斑马鱼的所有免疫器官中都有表达。我们利用CRISPR/Cas9介导的基因敲除斑马鱼(irf3-KO)证明了Irf3对病毒性出血性败血症病毒(VHSV)的抗病毒免疫机制。在本研究中,我们使用了截短的Irf3蛋白(由缺失10 bp的irf3编码)进行进一步研究。与野生型Irf3蛋白斑马鱼(WT)相比,注射VHSV后,irf3-KO斑马鱼表现出剂量依赖性的高死亡率和早期死亡率,证实了Irf3的抗病毒活性。根据VHSV挑战时下游基因的表达分析结果,我们推断Irf3缺乏会严重影响ifnphi1和ifnphi2的表达。然而,在感染后5天(dpi),irf3-KO中ifnphi3的表达与WT相比没有显著变化,而irf7的转录在5dpi后在irf3-KO中显示出相当大的增加,这表明irf7控制着ifnphi3的表达。在3 dpi时,isg15、viperin、mxa和mxb的表达明显减少,这也支持了Irf3缺乏对感染早期抗病毒活性的影响。irf3-KO斑马鱼的死亡率高于WT斑马鱼可能是由于irf3-KO斑马鱼的免疫反应延迟导致炎症和组织损伤增加。我们的研究结果表明,Irf3通过调节关键的免疫信号分子和调控抗病毒免疫基因在斑马鱼的抗病毒免疫中发挥作用。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Functional characterization of irf3 against viral hemorrhagic septicemia virus infection using a CRISPR/Cas9-mediated zebrafish knockout model

Interferon regulatory factors (IRFs) are transcription factors involved in immune responses, such as pathogen response regulation, immune cell growth, and differentiation. IRFs are necessary for the synthesis of type I interferons through a signaling cascade when pathogen recognition receptors identify viral DNA or RNA. We discovered that irf3 is expressed in the early embryonic stages and in all immune organs of adult zebrafish. We demonstrated the antiviral immune mechanism of Irf3 against viral hemorrhagic septicemia virus (VHSV) using CRISPR/Cas9-mediated knockout zebrafish (irf3-KO). In this study, we used a truncated Irf3 protein, encoded by irf3 with a 10 bp deletion, for further investigation. Upon VHSV injection, irf3-KO zebrafish showed dose-dependent high and early mortality compared with zebrafish with the wild-type Irf3 protein (WT), confirming the antiviral activity of Irf3. Based on the results of expression analysis of downstream genes upon VHSV challenge, we inferred that Irf3 deficiency substantially affects the expression of ifnphi1 and ifnphi2. However, after 5 days post infection (dpi), ifnphi3 expression was not significantly altered in irf3-KO compared to that in WT, and irf7 transcription showed a considerable increase in irf3-KO after 5 dpi, indicating irf7's control over ifnphi3 expression. The significantly reduced expression of isg15, viperin, mxa, and mxb at 3 dpi also supported the effect of Irf3 deficiency on the antiviral activity in the early stage of infection. The higher mortality in irf3-KO zebrafish than in WT might be due to an increased inflammation and tissue damage that occurs in irf3-KO because of delayed immune response. Our results suggest that Irf3 plays a role in antiviral immunity of zebrafish by modulating critical immune signaling molecules and regulating antiviral immune genes.

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来源期刊
ACS Applied Bio Materials
ACS Applied Bio Materials Chemistry-Chemistry (all)
CiteScore
9.40
自引率
2.10%
发文量
464
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