Besse S, Vigneron M, Pichard E, Puvion-Dutilleul F
Laboratoire de Biologie et Ultrastructure du Noyau de l'UPR 272 CNRS, Villejuif, France.
Gene Expr. 1995;4(3):143-61.
The response of the cellular RNA processing machinery to herpes simplex virus type 1 (HSV-1) infection was studied at the ultrastructural level in HeLa cells and compared to the distribution of RNA polymerase II molecules and viral RNA. Immunogold labeling of RNA polymerase II molecules revealed that viral genome transcription was restricted to filaments in an intranuclear, virus-induced region. This region also contained viral RNAs as revealed by in situ hybridization of two biotinylated viral DNA probes: a probe encompassing a limited portion of the viral genome (the F fragment) and a probe for the total genome. In addition, the latter probe revealed large amounts of viral RNA within the clusters of interchromatin granules, intranuclear structures of normal cells that became enlarged during HSV-1 infection. Components of spliceosomes were localized by in situ hybridization with biotinylated U1 and U2 DNA probes. The large viral region contained only traces of U1 and U2 RNAs, probably because of the low frequency of splices of viral transcripts. The clusters of interchromatin granules, however, accumulated U1 and U2 RNAs with the same frequency as in noninfected cells. Poly(A) RNA was detected by in situ hybridization of a biotinylated poly(dT) probe. Some was present over the filaments of the virus-induced region but most was accumulated in the clusters of interchromatin granules. Our data suggest, therefore, that the clusters of interchromatin granules, in addition to their involvement in spliceosome component assembly, might also be a transient storage site for some families of viral mRNA, possibly a sorting site that regulates their migration.
在HeLa细胞中，从超微结构水平研究了细胞RNA加工机制对单纯疱疹病毒1型（HSV-1）感染的反应，并与RNA聚合酶II分子和病毒RNA的分布进行了比较。RNA聚合酶II分子的免疫金标记显示，病毒基因组转录局限于核内病毒诱导区域的细丝中。通过两种生物素化病毒DNA探针的原位杂交显示，该区域也含有病毒RNA：一种探针覆盖病毒基因组的有限部分（F片段），另一种探针针对整个基因组。此外，后一种探针显示在染色质间颗粒簇中有大量病毒RNA，染色质间颗粒是正常细胞的核内结构，在HSV-1感染期间会增大。通过与生物素化的U1和U2 DNA探针进行原位杂交来定位剪接体的成分。大的病毒区域仅含有微量的U1和U2 RNA，可能是因为病毒转录本的剪接频率较低。然而，染色质间颗粒簇积累U1和U2 RNA的频率与未感染细胞相同。通过生物素化聚（dT）探针的原位杂交检测到聚（A）RNA。一些存在于病毒诱导区域的细丝上，但大部分积累在染色质间颗粒簇中。因此，我们的数据表明，染色质间颗粒簇除了参与剪接体成分组装外，可能还是某些病毒mRNA家族的临时储存位点，可能是一个调节其迁移的分选位点。
