Regulation of transcription in vitro from herpes simplex virus genes.
Regulation of transcription in vitro from herpes simplex virus genes.
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单纯疱疹病毒基因的体外转录调控。
DOI：
10.1128/jvi.60.3.950-959.1986
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发表时间：
1986
影响因子：
5.4
通讯作者：
Beard,P
中科院分区：
文献类型：
--
作者：
Pizer,LI;Tedder,DG;Betz,JL;Wilcox,KW;Beard,P
通过使用从单纯疱疹病毒早期糖蛋白D基因、晚期糖蛋白C基因、晚期VP 5基因和即刻早期ICP 22基因切割的DNA作为模板进行体外转录测定。来自未感染的HeLa细胞的悬浮培养物的核提取物有效地合成了来自立即早期和延迟早期类基因的RNA。在较小程度上，提取物也使用从晚期基因切割的DNA作为模板。从受感染细胞制备的提取物中抑制了即刻早期基因的转录。通过凝胶电泳、转移至硝酸纤维素和用特异性抗体探测对感染细胞提取物中的蛋白质进行分析，证明存在病毒调节蛋白ICP 4。来自感染细胞的核提取物的色谱分级分离产生富含ICP 4的蛋白质的混合物（级分VIII）（S. W. Faber和K.W. Wilcox，Nucleic Acids Res.，14：6067-6083，1986）。向体外测定中添加组分VIII影响转录。根据测定中的DNA，观察到抑制或刺激作用。RNA合成的抑制被发现时，从立即早期基因的DNA被用作模板，和刺激被发现时，从早期或晚期基因的DNA被使用。
In vitro transcription assays were carried out by using as templates DNAs cut from the herpes simplex virus early glycoprotein D gene, the late glycoprotein C gene, the late VP5 gene, and the immediate-early ICP22 gene. Nuclear extracts from suspension cultures of uninfected HeLa cells effectively synthesized RNAs from genes of the immediate-early and delayed-early classes. To a lesser extent, the extracts also used DNAs cut from the late genes as templates. Transcription from the immediate-early gene was inhibited in extracts prepared from infected cells. Analysis of the proteins in infected-cell extracts by gel electrophoresis, transfer to nitrocellulose, and probing with specific antibody demonstrated the presence of the viral regulatory protein ICP4. Chromatographic fractionation of nuclear extract from infected cells yielded a mixture of proteins (fraction VIII) enriched in ICP4 (S.W. Faber and K.W. Wilcox, Nucleic Acids Res., 14:6067-6083, 1986). Addition of fraction VIII to the in vitro assay affected transcription. Depending on the DNA in the assay, an inhibitory or stimulatory effect was observed. Inhibition of RNA synthesis was found when DNA from the immediate-early gene was used as a template, and stimulation was found when DNA from the early or late gene was used.