Bichko V V, Taylor J M
Fox Chase Cancer Center, Philadelphia, Pennsylvania 19111-2497, USA.
J Virol. 1996 Nov;70(11):8064-70. doi: 10.1128/JVI.70.11.8064-8070.1996.
When the small form of the delta antigen (deltaAg-S) was expressed from a cDNA expression plasmid and subsequently detected by immunofluorescence, it was found localized to the nucleoli. However, if the cDNA was cotransfected with a cDNA expressing a mutated hepatitis delta virus (HDV) genome that could only replicate by using the deltaAg-S provided by the first plasmid, then most of the deltaAg-S was redistributed to the nucleoplasm, largely to specific discrete nucleoplasmic sites or speckles; this pattern was stable for at least 50 days after transfection. These speckles coincided with those detected with an antibody to SC35, an essential non-small nuclear ribonucleoprotein splicing factor. Others have shown that SC35 speckles correspond to active sites of DNA-directed transcription by RNA polymerase II and also of RNA processing. We also found, in contrast to the cotransfections with the mutant HDV and the deltaAg-S provided in trans, that cells transfected with wild-type HDV showed a variable pattern of staining. The SC35-like speckle pattern of accumulation of delta antigen deltaAg was maintained for only 6 days, after which the pattern began to change. By 18 days posttransfection, a variety of different deltaAg staining patterns were observed. This pattern of change occurs at a time when the large form of the delta antigen deltaAg-L appears and HDV RNA synthesis begins to shut down. Our studies therefore support the interpretation that HDV RNA and deltaAg-S accumulate at SC35 speckle sites in the nucleoplasm. We speculate that these may be the sites at which HDV RNA is transcribed by RNA polymerase II and/or sites of HDV RNA processing. Furthermore, when deltaAg-L, as well as other mutant deltaAg accumulate, the speckle association is disrupted, thereby stopping HDV RNA replication.
当从cDNA表达质粒中表达δ抗原的小形式（δAg-S）并随后通过免疫荧光检测时，发现其定位于核仁。然而，如果该cDNA与表达突变型丁型肝炎病毒（HDV）基因组的cDNA共转染，该突变型HDV基因组只能利用第一个质粒提供的δAg-S进行复制，那么大多数δAg-S会重新分布到核质中，主要是到特定的离散核质位点或斑点；这种模式在转染后至少50天内是稳定的。这些斑点与用抗SC35抗体检测到的斑点一致，SC35是一种必需的非小核核糖核蛋白剪接因子。其他人已经表明，SC35斑点对应于RNA聚合酶II进行DNA定向转录以及RNA加工的活性位点。我们还发现，与用突变型HDV和反式提供的δAg-S共转染相反，用野生型HDV转染的细胞显示出可变的染色模式。δ抗原δAg积累的SC35样斑点模式仅维持6天，之后模式开始改变。到转染后18天，观察到多种不同的δAg染色模式。这种变化模式发生在δ抗原大形式（δAg-L）出现且HDV RNA合成开始停止的时候。因此，我们的研究支持这样的解释，即HDV RNA和δAg-S在核质中的SC35斑点位点积累。我们推测这些可能是HDV RNA由RNA聚合酶II转录的位点和/或HDV RNA加工的位点。此外，当δAg-L以及其他突变型δAg积累时，斑点关联被破坏，从而停止HDV RNA复制。
