ASSEMBLY OF ADENO-ASSOCIATED VIRUS
ASSEMBLY OF ADENO-ASSOCIATED VIRUS
复制标题
DOI：
10.1016/0042-6822(80)90071-9
复制
发表时间：
1980-01-01
期刊：
影响因子：
3.7
通讯作者：
CARTER, BJ
中科院分区：
文献类型：
--
作者：
MYERS, MW;CARTER, BJ
通过分别用[3 H]胸苷或[35 S]甲硫氨酸标记DNA和蛋白质前体，研究了缺陷细小病毒腺相关病毒（AAV）的组装。从细胞[人口腔癌KB]裂解物中提取病毒颗粒，并通过在CsCl梯度中平衡条带和在中性蔗糖中的速度沉降来表征。放射性脉冲追踪标记实验表明，新合成的AAV蛋白质快速组装成空衣壳，空衣壳反过来又充当含有标准或缺失（缺陷干扰）病毒基因组的稳定、成熟（完整）颗粒的前体。虽然新合成的子代DNA链与空衣壳的结合似乎是快速的，但成熟形成稳定的完整颗粒显然是一个慢得多的过程，需要几个小时。具有与处于DNA酶敏感构象的空衣壳相关的AAV基因组性质的新组分（与成熟颗粒中DNA的完全DNA酶抗性相反）也表现出与作为组装中间体的作用一致的脉冲追踪动力学。L-刀豆氨酸抑制空衣壳的产生，从而抑制完整颗粒的组装。感染的培养物与羟基脲预处理抑制外观的完整的颗粒，并导致空衣壳的积累与其前体的作用一致。在脉冲标记后立即添加羟基脲显示，AAV颗粒的成熟似乎在不存在伴随的DNA复制的情况下发生。提出了一个模型，其中子代DNA链包装成预制衣壳的AAV组装。
The assembly of the defective parvovirus, adeno-associated virus (AAV), was studied by labeling DNA and protein precursors with [3H]thymidine or [35S]methionine, respectively. Virus particles were extracted from cell [human oral carcinoma KB] lysates and characterized by banding to equilibrium in CsCl gradients and by velocity sedimentation in neutral sucrose. Radioactive pulse-chase labeling experiments showed that newly synthesized AAV proteins are rapidly assembled into empty capsids which in turn serve as precursors to stable, mature (full) particles that contain a standard or deleted (defective-interfering) virus genome. While association of the newly synthesized progeny DNA strands with empty capsids appears to be rapid, maturation to form a stable full particle apparently is a much slower process requiring several hours. A new component having the properties of an AAV genome associated with the empty capsid in a DNase-sensitive conformation (in contrast to the complete DNase resistance of DNA in mature particles) also exhibited pulse-chase kinetics consistent with a role as an assembly intermediate. L-Canavanine inhibited production of empty capsids and thus inhibited assembly of full particles. Pretreatment of infected cultures with hydroxyurea inhibited appearance of full particles and caused accumulation of empty capsids consistent with their precursor role. Addition of hydroxyurea immediately after a pulse label showed that maturation of AAV particles appears to occur in the absence of concomitant DNA replication. A model is proposed for AAV assembly in which progeny DNA strands are packaged into preformed capsids.