Cunha C, Monjardino J, Cheng D, Krause S, Carmo-Fonseca M
Institute of Histology and Embryology, Faculty of Medicine, University of Lisbon, Portugal.
RNA. 1998 Jun;4(6):680-93. doi: 10.1017/s135583829898013x.
Hepatitis delta virus (HDV) is a human pathogen that can greatly increase the severity of liver damage caused by an hepatitis B infection. HDV contains a circular, single-stranded RNA genome that encodes a unique protein, the delta antigen. Two forms of the delta antigen, deltaAg-S and deltaAg-L, are derived from a single open reading frame by RNA editing. Here we analyze the subcellular distribution of HDV RNA and its spatial relationship to known intranuclear structures. The human hepatoma cell line Huh7 was stably transfected with wild-type HDV cDNA and the viral RNAs were localized by in situ hybridization and fluorescence confocal microscopy. HDV RNA is detected throughout the nucleoplasm, with additional concentration in focal structures closely associated with nuclear speckles or clusters of interchromatin granules. Both the smaller form of the delta antigen (deltaAg-S), which is required for HDV genomic replication, and the larger form of the delta antigen (deltaAg-L), which represses replication, co-localize with delta RNA throughout the nucleoplasm and in the foci. However, the foci do not incorporate bromo-UTP and do not concentrate either RNA polymerase II or cleavage and polyadenylation factors required for viral RNA synthesis and 3' end processing, respectively. Thus, it is unlikely that the delta foci represent major sites of viral transcription or replication. In conclusion, the data show that viral RNA-protein complexes accumulate in structures closely associated with interchromatin granules, a subnuclear domain highly enriched in small nuclear ribonucleoproteins, poly(A+) RNA, and RNA splicing protein factors. This implies a specific compartmentalization of ribonucleoproteins in the nucleus.
丁型肝炎病毒（HDV）是一种人类病原体，可显著加重由乙型肝炎感染所导致的肝损伤的严重程度。HDV含有一个环状单链RNA基因组，该基因组编码一种独特的蛋白质，即δ抗原。δ抗原的两种形式，δAg-S和δAg-L，是通过RNA编辑从单一开放阅读框衍生而来。在此，我们分析了HDV RNA的亚细胞分布及其与已知核内结构的空间关系。用野生型HDV cDNA稳定转染人肝癌细胞系Huh7，并通过原位杂交和荧光共聚焦显微镜对病毒RNA进行定位。HDV RNA在整个核质中均有检测到，在与核斑点或染色质间颗粒簇紧密相关的局灶性结构中浓度更高。HDV基因组复制所需的较小形式的δ抗原（δAg-S）和抑制复制的较大形式的δ抗原（δAg-L），均与δRNA在整个核质和这些病灶中共定位。然而，这些病灶不掺入溴尿嘧啶三磷酸，也不富集病毒RNA合成及3'端加工分别所需的RNA聚合酶II或切割和聚腺苷酸化因子。因此，δ病灶不太可能代表病毒转录或复制的主要位点。总之，数据表明病毒RNA-蛋白质复合物在与染色质间颗粒紧密相关的结构中积累，染色质间颗粒是一个富含小核核糖核蛋白、多聚腺苷酸（A+）RNA和RNA剪接蛋白因子的核内亚结构域。这意味着核糖核蛋白在细胞核中存在特定的区室化。
