Cloning and structural analysis of four genes encoding interferon-omega in rabbit.

M Charlier, R L'Haridon, M Boisnard, J Martal, P Gaye
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引用次数: 16

Abstract

By using an ovine interferon-tau (IFN-tau) cDNA probe, four recombinant phages were isolated from a rabbit genomic library and sequenced from nucleotides -450 to 1,300 relative to the CAP site. Each of the four rabbit genes contains an open reading frame of 595 nucleotides and code for proteins that exhibit structural characteristics of the interferon-omega (IFN-omega) family. They display more than 98% identity in their coding regions. The deduced amino acid sequences share > 96% sequence similarity. In contrast, the 5' and 3' noncoding regions have diverged considerably (approximately 50% identity). Amino acid comparisons of rabbit IFN-omega with IFN-omega of other species reveal the highest degree of identity with human (72%), followed by porcine (68%) IFN-omega. Rabbit IFN-omega displays only 57% sequence similarity with ovine IFN-tau. The coding regions of the four genes subcloned in a cytomegalovirus eukaryotic expression vector and transfected in monkey COS-7 cells direct the production of proteins that protect bovine and rabbit cells against vesicular stomatitis virus infection, thus demonstrating that these genes encode fully active IFN proteins. The expression of these genes was studied in Sendai-induced rabbit leukocytes. A single band of poly(A)+RNA hybridized with a rabbit IFN-omega probe under stringent conditions, whereas no IFN-omega transcript was detected with RNA isolated from uninduced leukocytes. Southern blot analysis suggest the existence of at least eight IFN-omega genes or pseudogenes in the rabbit genome.

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兔干扰素编码基因的克隆及结构分析。
利用羊干扰素-tau (IFN-tau) cDNA探针,从兔基因组文库中分离到4个重组噬菌体,并对相对于CAP位点的-450 ~ 1300个核苷酸进行了测序。这四个兔子基因中的每一个都包含一个595个核苷酸的开放阅读框,并编码干扰素-omega (IFN-omega)家族结构特征的蛋白质。它们在编码区显示出98%以上的同一性。推导出的氨基酸序列相似性> 96%。相比之下,5'和3'非编码区差异很大(大约50%相同)。兔IFN-omega与其他物种的IFN-omega氨基酸比较显示,与人类IFN-omega的同源度最高(72%),其次是猪IFN-omega(68%)。兔IFN-omega与羊IFN-tau序列相似性仅为57%。在巨细胞病毒真核表达载体中亚克隆并转染猴COS-7细胞的4个基因的编码区可指导牛和兔细胞免受水泡性口炎病毒感染的蛋白的产生,从而证明这些基因编码完全活性的IFN蛋白。研究了这些基因在仙台诱导兔白细胞中的表达。在严格的条件下,单带poly(A)+RNA与兔IFN-omega探针杂交,而从未诱导的白细胞中分离的RNA未检测到IFN-omega转录物。Southern blot分析表明,家兔基因组中至少存在8个IFN-omega基因或假基因。
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