乙型肝炎病毒剪接变异与对干扰素治疗受损响应相关联

StephenW

Hepatitis B virus spliced variants are associated with an impaired response to interferon therapy

• 
  Jieliang Chen
• , Min Wu
• , Fan Wang
• , Wen Zhang
• , Wei Wang
• , Xiaonan Zhang
• , Jiming Zhang
• , Yinghui Liu
• , Yi Liu
• , Yanling Feng
• , Ye Zheng
• , Yunwen Hu
• & Zhenghong Yuan
  

• Scientific Reports 5, Article number: 16459 (2015)
• doi:10.1038/srep16459
• Download Citation
• 
  
  • Hepatitis B |
  • Hepatitis B virus |
  • Immune evasion |
  • Interferons
    

Received:04 August 2015Accepted:14 October 2015Published online:20 November 2015


AbstractDuring hepatitis B virus (HBV) replication, spliced HBV genomes and splice-generated proteins have been widely described, however, their biological and clinical significance remains to be defined. Here, an elevation of the proportion of HBV spliced variants in the sera of patients with chronic hepatitis B (CHB) is shown to correlate with an impaired respond to interferon-α (IFN-α) therapy. Transfection of the constructs encoding the three most dominant species of spliced variants into cells or ectopic expression of the two major spliced protein including HBSP and N-terminal-truncated viral polymerase protein result in strong suppression of IFN-α signaling transduction, while mutation of the major splicing-related sites of HBV attenuates the viral anti-IFN activities in both cell and mouse models. These results have associated the productions of HBV spliced variants with the failure response to IFN therapy and illuminate a novel mechanism where spliced viral products are employed to resist IFN-mediated host defense.


IntroductionHepatitis B virus (HBV), which can cause chronic liver disease that may result in cirrhosis and hepatocellular carcinoma (HCC), is a DNA virus that has a 3.2 kB partially double-stranded, relaxed circular genome organized into overlapping open-reading frames from which viral genes are transcribed. The major HBV messenger RNAs (mRNA) synthesized from the HBV genome include: 3.5 kb (long/short) preC/pre-genomic (pg) RNA that encodes hepatitis B e antigen (HBeAg), core and polymerase (Pol) proteins, 2.4 kb and 2.1 kb sub-genomic RNAs that encode the HBV surface proteins (HBs), and a 0.9 kb mRNA that encodes the HBx protein. Through the 5′ stem-loop structure ε, the 3.5 kb pgRNA can also be packaged and reverse-transcribed by the viral polymerase into relaxed circular DNA to generate the wild-type HBV (wtHBV) particles for secretion into circulation1.
In addition to the unspliced mRNAs, a series of spliced (SP) HBV (spHBV) RNAs have been widely described in model systems and in HBV-infected livers2,3,4,5. The most frequently detected spHBV variant is a 2.2 kb molecule termed SP1, which is generated through the removal of a 1.3 kb intron from the pgRNA and accounts for up to 30% of pgRNAs3,4,5,6,7,8. spHBV RNAs can be incorporated into the nucleocapsids and then reverse transcribed into HBV DNA to generate defective HBV (dHBV) particles9,10,11,12. The level of dHBV particles in the sera of patients with chronic hepatitis B (CHB) was shown to be related with liver disease13,14 and was enhanced prior to development of HCC15. spHBV RNAs also serve as the translation templates for a number of non-canonical HBV proteins. HBV splice-generated protein (HBSP) is one of the major spHBV RNAs-encoded proteins which was firstly identified by Soussan et al. in the livers of CHB patients16, and has been reported to be associated with viral replication, liver diseases and cancer development17,18,19. In spite of these evidences suggesting that HBV splicing is a common event during HBV infection and may be involved in the pathogenicity or persistence of HBV, the clinical and biological significance of HBV splicing needs to be further defined.
Interferon-α (IFN-α), a cytokine with antiviral and immunomodulatory activities, has been approved for treatment of HBV infection since 1990s. However, IFN therapy is effective in only 30–40% of CHB patients20,21. One possible explanation for the failure to respond to IFN therapy in CHB patients is that HBV has developed strategies to counteract the IFN signaling transduction22,23,24,25,26,27. The Pol protein consisting of terminal protein (TP), reverse transcriptase (RT), RNaseH (RH), and a non-conserved spacer domain between the TP and RT domains has been shown to be multifunctional, not only playing essential role during viral replication but also having modulatory functions by interacting with a number of host factors28. Foster et al. found that the expression of the TP region of Pol inhibits cellular responses to IFN-α22,23, and we further observed that the TP and RH domains of Pol inhibit IFN-activated STAT1 serine 727 phosphorylation and STAT1 and STAT2 nuclear translocation, respectively25. Since Pol is believed to be produced at a relatively low level during viral replication, the real physiological relevance of these findings remains to be classified.
Considering that most of the spHBV variants contain sequences that could be translated into proteins which contain domains with anti-IFN activities derived from the open reading frame (ORF) of Pol gene11,29, we hypothesized that there may be a correlation between HBV spliced variants and IFN responses. To test this hypothesis, we collected serum samples from CHB patients before and after IFN therapy to analyze the relationship between the productions of HBV spliced variants and the failure to respond to IFN therapy. In addition, we examined the expression of HBV spliced variants and investigated the potential role of HBV splicing in counteraction of IFN signaling in cell culture systems and in a mouse model.

StephenW

乙型肝炎病毒剪接变异与对干扰素治疗受损响应相关联

    姐俩陈，吴敏，王帆，张文，王炜，张小囡，继铭张应辉刘，刘怡，炎陵枫，叶征，马蕴雯胡锦涛和正虹元

    科学报告5，商品编号：16459（2015年）
    DOI：10.1038 / srep16459
    下载文献
        乙肝|乙肝病毒|免疫逃避|干扰素

收稿日期：
    2015年08月04日
公认：
    2015年10月14日
网络发布时间：
    2015年11月20日

抽象

期间乙型肝炎病毒（HBV）的复制，剪接的HBV基因组和剪接生成的蛋白质已经被广泛地描述的，然而，它们的生物和临床意义仍有待确定。这里，乙型肝炎病毒剪接变体在治疗慢性乙型肝炎（CHB）的血清的比例的正视被证实与对干扰素α（IFN-α）治疗受损的响应。转染剪接变体的三个最优势种编码进入细胞或两个主要剪接蛋白包括HBSP和N-末端截短的病毒聚合酶蛋白导致IFN-α信号转导的强有力的抑制的异位表达的构建物，而该突变乙肝病毒主要剪接相关网站减弱在细胞和小鼠模型的抗病毒干扰素的活动。这些结果相关联的失败响应干扰素治疗HBV剪接变异体的生产和照亮的地方拼接病毒产品被用来抵抗干扰素介导的宿主防御新机制。
介绍

乙型肝炎病毒（HBV），这可引起慢性肝脏疾病可能导致肝硬化和肝细胞癌（HCC）的是，是一种DNA病毒，其具有3.2kb的部分双链，松弛环状组织成重叠从开放阅读框的基因组该病毒基因转录。从HBV基因合成的主要乙肝病毒信使RNA（mRNA）的包括：3.5 KB（长/短）前C /预基因组（PG）RNA编码乙型肝炎e抗原（HBeAg），核心和聚合酶（POL）的蛋白质，2.4 kb和2.1kb的编码HBV表面蛋白（HBs）中和亚基因组RNA的，和一个0.9kb的mRNA的编码该HBx蛋白。通过5'茎环结构ε，该3.5kb的pgRNA也可包装和反转录的病毒聚合酶成放宽环状DNA，以产生野生型HBV（wtHBV）颗粒分泌到circulation1。

除了未剪接的mRNA，一系列剪接（SP）的HBV（spHBV）的RNA已广泛在模型系统中和在HBV感染livers2,3,4,5说明。最常检测到的spHBV变体是一个2.2kb的分子称为SP 1，这是通过移除从pgRNA一个1.3kb的内含子的生成和占pgRNAs3,4,5,6,7,8的高达30％。 spHBV的RNA可以被掺入到核衣壳，然后反转录成的HBV DNA，以产生有缺陷的HBV（DHBV）particles9,10,11,12。 DHBV颗粒在患有慢性乙型肝炎（CHB）的血清水平被证明与肝disease13,14有关和增强之前HCC15的发展。 spHBV的RNA也可以作为翻译模板的一些非规范的HBV蛋白的。 HBV的拼接产生的蛋白质（HBSP）是将其首先通过Soussan等人确定的主要spHBV的RNA编码的蛋白之一。在CHB patients16的肝脏，并且已经报道与病毒复制，肝脏疾病和癌症development17,18,19相关联。尽管这些证据表明HBV剪接是HBV感染期间常见事件，并可能参与的HBV的致病性或持续性的，需要进一步定义的HBV剪接的临床和生物学意义。

干扰素α（IFN-α），具有抗病毒和免疫调节活性的细胞因子，已被批准用于1990年以来治疗HBV感染。然而，IFN治疗是有效的仅30-40％的CHB patients20,21的。一个可能的解释未能在CHB患者对干扰素治疗是乙肝病毒已经发展战略，以抵消IFN信号transduction22,23,24,25,26,27。的聚合酶蛋白选自由末端蛋白（TP）的，逆转录酶（RT），RNA酶H（RH）中，并且在TP和RT域之间的非保守间隔结构域已被证明是多功能的，而不是病毒复制过程中只扮演重要的角色而且通过与一些宿主factors28的相互作用具有调节功能。 Foster等。发现，波尔的TP区域的表达抑制细胞应答于IFN-α22,23，我们还观察到的波尔抑制干扰素活化的STAT1丝氨酸727的磷酸化和STAT1和STAT2核易位，respectively25在TP和RH的域。因为POL被认为是在病毒复制过程中相对较低的水平产生，在这些发现的真实生理相关仍有待分类。

考虑到大多数的spHBV变体包含这样的序列可以转化成包含结构域，从开放读码聚合酶gene11,29的（ORF）衍生的抗IFN活性的蛋白质，我们推测，可能存在的HBV剪接变体之间的相关性和IFN的反应。为了检验这一假设，我们收集从慢性乙型肝炎患者的血清样品之前和IFN治疗后分析的HBV变体剪接作品以及未能对IFN治疗的反应之间的关系。另外，我们研究了HBV的剪接变体的表达和研究中的IFN信号传导在细胞培养系统的反作用，并在小鼠模型中的HBV剪接的潜在作用。

StephenW

全文:http://www.nature.com/articles/s ... rtId=ODAzMjY0Nzk4S0

生命满希望

什么意思？什么变异，受损？

StephenW

回复 生命满希望 的帖子

生物学非常复杂.

選擇性剪接（英语：Alternative splicing；又称“可变剪接”）是基因表达的方式，在人類细胞是非常普遍的。真核细胞的基因序列中，包含了內含子（intron）與外顯子（exon），兩者交互穿插。其中內含子在基因轉錄成mRNA前体後會被RNA剪接體移除，剩下的外顯子才是能夠存在於成熟mRNA（之後再進一步轉譯成蛋白質）的片段。

而選擇性剪接便是利用這樣的特性，一條未經剪接的RNA，含有的多種外顯子被剪成的不同組合，可轉譯出不同的蛋白質。就能將同一基因中的外顯子以不同的組合方式來表現，使一個基因在不同時間、不同環境中能夠製造出不同的蛋白質（基因表达调控），可增加生理狀況下系統的複雜性或適應性。