Lei Yan-Chang, Ma Tao, Hao You-Hua, Zhang Zheng-Mao, Tian Yong-Jun, Wang Bao-Ju, Yang Dong-Liang
Division of Clinical Immunology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
Zhonghua Gan Zang Bing Za Zhi. 2006 Oct;14(10):738-41.
To investigate the effect of apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3G (APOBEC3G) mediated antiviral activity against hepatitis B virus (HBV) and duck hepatitis B virus (DHBV).
Total RNA was extracted from peripheral blood mononuclear cells (PBMCs), RT-PCR product was cloned into the EcoR I/Hind III restriction sites of the CMV-driven expression vector fused with a hemagglutinin fusion epitope tag at its carboxyl terminal. Replication competent 1.3 fold over-length HBV was constructed with full-length HBV of ayw subtype. The mammalian hepatoma cell HepG2 was cotransfected with the replication competent 1.3 fold over-length HBV and various amounts of CMV-driven expression vector encoding APOBEC3G-HA. Levels of HBsAg and HBeAg in the media of the transfected cells were determined by ELISA, HBV DNA. RNA from intracellular core particles was examined using Northern and Southern blot analyses. Chicken hepatoma cell LMH was cotransfected with head-to-tail dimer of an EcoR I monomer of DHBV and various amounts of CMV-driven expression vector encoding APOBEC3G-HA. DHBV DNA from intracellular core particles was examined using Southern blot analysis.
CMV-driven expression vector encoding APOBEC3G-HA and replication competent 1.3 fold over-length HBV were constructed. There was a dose dependent decrease in the levels of intracellular core-associated viral (HBV and DHBV) DNA and extracellular production of HBsAg and HBeAg. Levels of intracellular core-associated viral RNA were also decreased, but the expression of HBcAg remained almost unchanged.
APOBEC3G suppresses HBV and DHBV replication and also suppresses HBsAg and HBeAg expression.
研究载脂蛋白B mRNA编辑酶催化多肽样3G（APOBEC3G）介导的抗乙型肝炎病毒（HBV）和鸭乙型肝炎病毒（DHBV）的抗病毒活性。
从外周血单个核细胞（PBMC）中提取总RNA，将RT-PCR产物克隆到CMV驱动的表达载体的EcoR I/Hind III酶切位点，该载体在其羧基末端融合了血凝素融合表位标签。用ayw亚型的全长HBV构建具有复制能力的1.3倍超长HBV。将具有复制能力的1.3倍超长HBV与不同量的编码APOBEC3G-HA的CMV驱动表达载体共转染哺乳动物肝癌细胞HepG2。通过ELISA、HBV DNA检测转染细胞培养基中HBsAg和HBeAg的水平。使用Northern和Southern印迹分析检测细胞内核颗粒中的RNA。将DHBV的EcoR I单体的头对头二聚体与不同量的编码APOBEC3G-HA的CMV驱动表达载体共转染鸡肝癌细胞LMH。使用Southern印迹分析检测细胞内核颗粒中的DHBV DNA。
构建了编码APOBEC3G-HA的CMV驱动表达载体和具有复制能力的1.3倍超长HBV。细胞内核相关病毒（HBV和DHBV）DNA水平以及HBsAg和HBeAg的细胞外产生呈剂量依赖性降低。细胞内核相关病毒RNA水平也降低，但HBcAg的表达几乎保持不变。
APOBEC3G抑制HBV和DHBV复制，也抑制HBsAg和HBeAg表达。
