HBV RNA pre-genome encodes specific motifs that mediate interactions with the viral core protein that promote nucleocapsid assembly.
HBV RNA pre-genome encodes specific motifs that mediate interactions with the viral core protein that promote nucleocapsid assembly.
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DOI：
10.1038/nmicrobiol.2017.98
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发表时间：
2017-06-19
影响因子：
28.3
通讯作者：
Stockley PG
中科院分区：
文献类型：
--
作者：
Patel N;White SJ;Thompson RF;Bingham R;Weiß EU;Maskell DP;Zlotnick A;Dykeman E;Tuma R;Twarock R;Ranson NA;Stockley PG
B型肝炎病毒（HBV）核衣壳（NC）的形成是病毒生命周期中的重要步骤，但其组装尚未完全了解。我们报告发现的序列特异性之间的相互作用的病毒前基因组和HBV核心蛋白（Cp），在定义NC装配途径中发挥作用。使用RNA SELEX和生物信息学，我们在前基因组RNA中鉴定出对Cp二聚体具有高亲和力的多个区域。这些RNA形成具有保守环基序的茎环，其以比不存在RNA时高得多的保真度和产率触发病毒样颗粒（VLP）的序列特异性组装。RNA寡核苷酸不与预先形成的无RNA VLP相互作用，因此它们的作用必须在颗粒组装期间发生。在存在一种RNA的情况下组装的T=4 VLP的不对称冷冻-EM重建揭示了通过密度叶与主Cp壳连接的独特内部特征。生物物理测定表明，这是一个复杂的结构，涉及几个RNA寡聚体与Cp的C末端富含精氨酸的结构域相互作用。这些Cp-RNA接触可能在核衣壳组装期间调节前基因组的组织中发挥作用，促进随后的逆转录并充当NC组装的成核复合物。
Formation of the Hepatitis B (HBV) nucleocapsid (NC) is an essential step in the viral lifecycle but its assembly is not fully understood. We report the discovery of sequence-specific interactions between the viral pre-genome and HBV core protein (Cp) that play roles in defining the NC assembly pathway. Using RNA SELEX and bioinformatics we identified multiple regions in the pre-genomic RNA with high-affinity for Cp dimers. These RNAs form stem-loops with a conserved loop motif that trigger sequence-specific assembly of virus-like particles (VLPs) at much higher fidelity and yield than in the absence of RNA. The RNA oligos do not interact with preformed RNA-free VLPs, so their effects must occur during particle assembly. Asymmetric cryo-EM reconstruction of the T=4 VLPs assembled in the presence of one of the RNAs reveals a unique internal feature connected to the main Cp shell via lobes of density. Biophysical assays suggest that this is a complex involving several RNA oligos interacting with the C-terminal arginine-rich domains of Cp. These Cp-RNA contacts may play a role(s) in regulating the organization of the pre-genome during nucleocapsid assembly, facilitating subsequent reverse transcription and acting as a nucleation complex for NC assembly.