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鉴定和表达 ADAR1 基因,负责 RNA 编辑,以响应双链 RNA 和 GCRV 挑战在草鱼(Ctenopharyngodon idella)。

Identification and expression profiles of ADAR1 gene, responsible for RNA editing, in responses to dsRNA and GCRV challenge in grass carp (Ctenopharyngodon idella).

机构信息

Northwest A&F University, Shaanxi Key Laboratory of Molecular Biology for Agriculture, No. 22 Xinong Rd., Yangling 712100, China.

出版信息

Fish Shellfish Immunol. 2012 Oct;33(4):1042-9. doi: 10.1016/j.fsi.2012.07.002. Epub 2012 Jul 14.

DOI:10.1016/j.fsi.2012.07.002
PMID:22796906
Abstract

ADAR (adenosine deaminase acting on RNA) is an RNA editing enzyme that targets both coding and noncoding dsRNAs (double stranded RNAs) and converts adenosine to inosine, which is read by translation machinery and by polymerases during RNA-dependent RNA replication as if it is guanosine. This editing is a widespread post-transcriptional modification event in animals. In this study, we identified the full-length cDNA sequence of Ctenopharyngodon idella ADAR1 (designated as CiADAR1) and detected the mRNA expression patterns in response to dsRNA (polyinosinic-polycytidylic acid sodium salt, poly(I:C)) and grass carp reovirus (GCRV). CiADAR1 is a large gene in size, consisting of 4833 nucleotides encoding a protein of 1392 amino acids. The deduced amino acid sequence contains seven putative domains: one proline-rich region (Pro-R), two Z-DNA-binding domains (Zalpha), three dsRNA binding motifs (DSRM) and one tRNA-specific and dsRNA adenosine deaminase domain (ADEAMc). It is most homologous to Danio rerio ADAR (E-value = 0.0, identities = 80% (1110/1395)), also close homology to Homo sapiens ADAR1 (E-value = 0.0, identities = (47%) 530/1122). CiADAR1 mRNA was investigated in fifteen tissues, and was low expressed in muscle and head kidney tissues, high in blood and spleen tissues by quantitative real-time RT-PCR (qRT-PCR). mRNA expressions of CiADAR1 were significantly up-regulated and reached peak at 24 h post GCRV challenge in vivo and in vitro (P < 0.05). After poly(I:C) stimulation at different concentrations, CiADAR1 transcripts were rapidly and significantly up-regulated and recovered in dose-dependent and time-dependent manners (P < 0.05). The results indicate CiADAR1 was implicated in the antiviral immune response and laid the foundation for further studies on functions and mechanisms of RNA editing in fishes.

摘要

ADAR(腺苷脱氨酶作用于 RNA)是一种 RNA 编辑酶,可靶向编码和非编码双链 RNA(dsRNA),并将腺苷转化为肌苷,肌苷在翻译机制和依赖 RNA 的 RNA 复制期间被聚合酶读取,就像它是鸟苷一样。这种编辑是动物中广泛存在的转录后修饰事件。在这项研究中,我们鉴定了草鱼 ADAR1 的全长 cDNA 序列(命名为 CiADAR1),并检测了其对 dsRNA(聚肌苷-聚胞苷酸钠盐,poly(I:C)) 和草鱼虹彩病毒(GCRV)的 mRNA 表达模式。CiADAR1 是一个较大的基因,由 4833 个核苷酸组成,编码 1392 个氨基酸的蛋白质。推导的氨基酸序列包含七个假定的结构域:一个脯氨酸丰富区(Pro-R)、两个 Z-DNA 结合结构域(Zalpha)、三个 dsRNA 结合基序(DSRM)和一个 tRNA 特异性和 dsRNA 腺苷脱氨酶结构域(ADEAMc)。它与斑马鱼 ADAR 最为同源(E 值=0.0,身份=80%(1110/1395)),与人类 ADAR1 也有密切的同源性(E 值=0.0,身份=47%(530/1122))。通过定量实时 RT-PCR(qRT-PCR)检测 CiADAR1 在十五种组织中的表达情况,在肌肉和头肾组织中低表达,在血液和脾脏组织中高表达。体内和体外 GCRV 攻毒后 24 小时 CiADAR1 的 mRNA 表达显著上调并达到峰值(P<0.05)。用不同浓度的 poly(I:C)刺激后,CiADAR1 转录物迅速且显著上调,并呈剂量和时间依赖性恢复(P<0.05)。结果表明 CiADAR1 参与了抗病毒免疫反应,为进一步研究鱼类 RNA 编辑的功能和机制奠定了基础。

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