水牛中与疯牛病易感相关的朊病毒基因多态性研究

发布时间：2018-03-11 01:34

本文选题：朊病毒基因　切入点：多态性　出处：《云南大学》2015年硕士论文　论文类型：学位论文

【摘要】：朊病毒疾病,又名传染性海绵状脑病,是一类具有高度致死性和传染性的神经退行性疾病。该类疾病已在十几种哺乳动物中发现,例如人类的克雅氏症、羊的瘙痒病以及牛的疯牛病等,给人类和动物健康带来了严重威胁,同时给社会经济造成了巨大损失,因此,对朊病毒疾病的研究已经成为国内外医学和生物学领域研究的热点。朊病毒疾病的致病因子是朊蛋白(prion protein, PrP),它由朊病毒基因(prion protein gene, PRNP)编码,在朊病毒疾病的致病机制中起到至关重要的作用。已有的研究证实包括人在内的动物对朊病毒疾病的易感性与PRNP基因的多态性相关。其中,与疯牛病易感性相关的PRNP基因多态性包括以下四个方面：1)位于告PRNP基因启动子区域的23-bp插入/缺失(insertion/deletion,indel)多态；2)位于牛PRNP基因内含子1的12-bp indel多态；3)牛PRNP基因编码区的寡肽重复数目；4)牛PRNP基因编码区的氨基酸多态性。令人注目的是,到目前为止全世界已有超过19万头的黄牛被报道感染疯牛病,尽管黄牛和水牛的系统发育关系很相近,但是没有一头水牛被报道感染疯牛病(OIE统计)。之前的研究都主要集中在对疯牛病易感的黄牛上,而对疯牛病不易感的水牛PRNP基因遗传多样性的研究甚少。在本研究中,我们调查了涵盖8个不同品种的312头中国水牛的PRNP基因中四种多态的频率分布情况,包括水牛PRNP基因23-bp indel多态、12-bp indel多态、开放阅读框中寡肽重复数多态以及编码区的氨基酸多态。然后,汇集文献中已经报道过的所有患疯牛病的黄牛、健康黄牛以及水牛群体的23-bp和12-bp indel多态的数据进行全面地比较分析,我们的研究有三个重要的发现：1)在水牛PRNP基因23-bp和12-bp indel多态中,与疯牛病易感性密切相关的缺失型等位基因(D23和D12)频率非常低；2)水牛PRNP基因编码区的6个寡肽重复的等位基因频率显著低于黄牛；3)水牛编码区中有7个位点发生了非同义突变,且在两个物种中的频率分布有显著差异：S4R、A16V、P54S、G108S、V123M、S154N和F257L,其中,S4R、A16V和V123M为种间固定突变位点,水牛的54S、154N和257L三个位点的等位基因是固定的,而在黄牛中的频率分布却非常低。相反的,在黄牛中G108位点是固定的,而在93.1%的水牛中是丝氨酸,其余的是甘氨酸。另外,基于PRNP编码区的研究结果,以及在疯牛病中PrP发生蛋白构象转换及倾向发生聚集这一分子特征,我们对野生型的黄牛PrP和水牛中含有的突变型PrP(S154N)进行分子动力学模拟分析,分析的结果表明突变能够弱化聚集趋势和构象转化的结论。因此,PrP分子动力学模拟结果显示黄牛比水牛更易于发生构象转变,这可能与黄牛易感染疯牛病相关。这些研究结果表明牛PRNP基因的遗传差异可能与物种对疯牛病的易感性相关。我们的发现将有助于解释这两种关系相近的动物对疯牛病易感性的差异。
[Abstract]:Prion disease, also known as infectious spongiform encephalopathy, is a highly lethal and infectious neurodegenerative disease that has been found in more than a dozen mammals, such as Creutzfeldt-Jakob disease in humans. The pruritus of sheep and mad cow disease of cattle have brought serious threat to human and animal health, at the same time have caused huge losses to social economy, so, The research of prion disease has become a hot spot in the field of medicine and biology at home and abroad. The cause of prion disease is prion protein, PRPX, which is encoded by the prion gene prion protein genetics (PRNPN). Prion disease plays a crucial role in the pathogenesis of prion disease. Previous studies have shown that prion disease susceptibility in animals, including humans, is associated with polymorphism in the PRNP gene. The polymorphism of PRNP gene associated with susceptibility to mad cow disease includes the following four aspects: 1) 23-bp insertion / deletion in the promoter region of PRNP gene insertion / deletion in del2) 12-bp indel polymorphism in bovine PRNP gene intron 1) bovine PRNP gene coding region. Amino acid polymorphism in the coding region of bovine PRNP gene. More than 190,000 cattle around the world have been reported to be infected with mad cow disease so far, although the phylogenetic relationship between cattle and buffalo is very similar. However, no buffalo was reported to be infected with mad cow disease. Previous studies focused on cattle susceptible to mad cow disease, but few studies were conducted on the genetic diversity of PRNP gene in buffalo that was not susceptible to mad cow disease. In this study, we investigated the frequency distribution of four polymorphisms in the PRNP gene of 312 Chinese buffaloes covering 8 different breeds, including the 23-bp indel polymorphism of the PRNP gene of buffalo and 12-bp indel polymorphism. Oligoseptide repeat polymorphisms and amino acid polymorphisms in the coding region in the open reading box. Then, all the cattle with mad cow disease reported in the literature were collected. The data of 23-bp and 12-BP indel polymorphisms in healthy cattle and buffalo populations were compared and analyzed. Three important findings of our study were found in the PRNP gene 23-bp and 12-BP indel polymorphisms of buffalo. The frequency of deletion alleles (D23 and D12), which are closely related to the susceptibility of mad cow disease, is very low. The allele frequency of 6 oligoseptide repeats in the PRNP coding region of buffalo is significantly lower than that in the buffalo coding region, and there are 7 non-synonymous mutations in the coding region of buffalo. There were significant differences in the frequency distribution between the two species. There were significant differences in the frequency distribution between the two species. There were significant differences in the frequency distribution between the two species. Among them, S4RNA16V and V123M were the interspecific fixed mutation sites, and the alleles of 54Sn154N and 257L were fixed in buffalo, but the frequency distribution in yellow cattle was very low. The G108 locus was fixed in yellow cattle, serine in 93.1% buffalo and glycine in the rest. In addition, based on the results of PRNP coding region, and the molecular characteristics of protein conformation conversion and tendency of aggregation of PrP in mad cow disease, We simulated the molecular dynamics of wild type yellow cattle PrP and buffalo mutant PrPS154N. The results of analysis showed that the mutation could weaken the conformation transformation and the aggregation trend. Therefore, the results of PrP molecular dynamics simulation showed that yellow cattle were more prone to conformation transition than buffalo. These results suggest that genetic differences in bovine PRNP gene may be associated with the species susceptibility to mad cow disease. Our findings will help explain the relationship between the two closely related animals. The difference in susceptibility to mad cow disease.
【学位授予单位】：云南大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S852.653

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