Hepatitis B virus X protein (HBx)-induced apoptosis in HuH-7 cells: Influence of HBV genotype and basal core promoter mutations
Hepatitis B virus X protein (HBx)-induced apoptosis in HuH-7 cells: Influence of HBV genotype and basal core promoter mutations
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DOI：
10.1080/00365520310008719
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发表时间：
2004-05-01
影响因子：
1.9
通讯作者：
Saisho, H
中科院分区：
文献类型：
--
作者：
Kanda, T;Yokosuka, O;Saisho, H
背景：B型肝炎病毒（HBV）感染是一个严重的世界性问题。HBV基因型和基础核心启动子（BCP）突变影响HBV感染患者的临床病程。BCP突变还导致HBV X蛋白（HBx）密码子130/131处的突变。人HBx天然变异体的功能意义在很大程度上仍不清楚。本研究的目的是探讨HBV基因型或双突变是否影响HBx诱导的细胞凋亡。研究方法：我们构建了基因型A、B、C和D的HBx表达载体和使用定点突变在HBx密码子130 K和131 V或位置130 M和131 I处具有双突变的HBx表达载体。建立了HuH-7细胞中的瞬时表达系统，并利用该模型来解决HBx对细胞活力的影响。结果：MTS检测显示HBx转染的细胞活力呈剂量依赖性降低。根据末端转移酶介导的DNA末端标记和caspase-3活性，表达HBx的细胞亚群发生凋亡。这项研究表明，HBx可以诱导细胞死亡的细胞凋亡，在剂量依赖性的方式和HBV基因型和双突变不影响HBx诱导的细胞凋亡。结论：HBV基因型和两个氨基酸的突变直接相邻的保守Kunitz结构域的转录激活活性的HBx没有改变HBx的促凋亡活性。需要进一步研究以确定HBV基因型和双突变是否对HBx的功能有任何影响。
Background: Hepatitis B virus (HBV) infection is a serious, world-wide problem. HBV genotype and basal core promoter (BCP) mutations affect the clinical course of HBV-infected patients. BCP mutations also lead to mutations at HBV X protein (HBx) codons 130/131. The functional significance of naturally occurring variants of human HBx remains largely unknown. The purpose of the study was to investigate whether HBV genotypes or double mutations affect HBx-induced apoptosis. Methods: We constructed genotype A, B, C, and D HBx expression vectors and HBx expression vectors with double mutations at HBx codons 130K and 131V or positions 130M and 131I using site-directed mutagenesis. A transient expression system in HuH-7 cells was established and this model was utilized to address the effect of HBx on cell viability. Results: HBx-transfected cells showed a dose-dependent decrease in cell viability by MTS assay. A subset of cells expressing HBx underwent apoptosis according to terminal transferase enzyme-mediated end labeling of DNA and caspase-3 activity. This study demonstrated that HBx can induce cell death by apoptosis in a dose-dependent manner and that HBV genotypes and double mutations did not affect HBx-induced apoptosis. Conclusions: HBV genotypes and mutation of two amino acids directly adjacent to the conserved Kunitz domain essential for transcription activating activity of HBx did not change the pro-apoptotic activity of HBx. Further study is needed to determine whether HBV genotypes and double mutations have any effect on the function of HBx.