Differential activation of hybrid genes containing herpes simplex virus immediate-early or delayed-early promoters after superinfection of stable DNA-transfected cell lines.
Differential activation of hybrid genes containing herpes simplex virus immediate-early or delayed-early promoters after superinfection of stable DNA-transfected cell lines.
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稳定 DNA 转染细胞系重复感染后，含有单纯疱疹病毒立即早期或延迟早期启动子的杂交基因的差异激活。
DOI：
10.1128/jvi.56.3.867-878.1985
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发表时间：
1985
影响因子：
5.4
通讯作者：
Hayward,GS
中科院分区：
文献类型：
--
作者：
Mosca,JD;Reyes,GR;Pitha,PM;Hayward,GS
我们比较了单纯疱疹病毒（HSV）在包含人β -干扰素（IFN）或氯霉素乙酰转移酶（CAT）基因的细胞系在HSV立即早期（IE）或延迟早期启动子控制下的重复感染后获得的基因表达水平。dna转染的小鼠Ltk+细胞系含有共同选择的IE175-IFN或胸苷激酶(TK)-IFN杂交基因，只有低水平的人IFN基础表达。然而，HSV 1型或HSV 2型感染两种细胞类型都会产生大量的IFN特异性RNA合成和具有生物活性的IFN蛋白产物。IE175-IFN细胞系的IFN滴度持续增加20- 150倍，一些TK-IFN细胞系的诱导度达到100- 500倍。在IE175-IFN细胞中，IFN RNA的表达也增加了200倍，并在病毒感染后30 - 60分钟内检测到。在含有共选择的IE175-CAT和TK-CAT构建体的抗g418杂交Ltk-或Vero细胞系中，除了IE175-CAT的基础表达相对较高外，也获得了类似的定性结果。所有三组IE细胞系（但不包括延迟-早期细胞系）对环己亚胺存在和IE基因表达缺陷突变的病毒感染都有反应，表明IE前病毒粒子因子特异性反式激活。相比之下，TK杂交细胞类型的激活需要病毒基因表达和功能性IE175基因产物的存在。在IE175杂交细胞感染HSV后的20小时内，驻留的IFN或CAT DNA序列的拷贝数也出现了高达30倍的扩增，而在TK杂交细胞中则没有。用醋酸磷处理或用不能合成病毒DNA的ts突变体重复感染可消除扩增，这表明在IE杂交构建体中存在的病毒DNA复制起源（oriS）的特异性HSV激活。
We compared the levels of gene expression obtained after herpes simplex virus (HSV) superinfection of cell lines containing integrated human beta-interferon (IFN) or chloramphenicol acetyltransferase (CAT) genes under the control of HSV immediate-early (IE) or delayed-early class promoters. DNA-transfected mouse Ltk+ cell lines harboring coselected IE175-IFN or thymidine kinase (TK)-IFN hybrid genes gave only low basal expression of human IFN. However, infection of both cell types with HSV type 1 or HSV type 2 produced abundant synthesis of IFN-specific RNA and biologically active IFN protein product. The IE175-IFN cell lines consistently gave 20- to 150-fold increases in IFN titers, and several TK-IFN cell lines yielded 100- to 500-fold induction. In the IE175-IFN cells, expression of IFN RNA also increased up to 200-fold and was detectable within 30 to 60 min after virus infection. Qualitatively similar results were obtained with hybrid G418-resistant Ltk- or Vero cell lines containing coselected IE175-CAT and TK-CAT constructs, except that there was relatively high basal expression of IE175-CAT. All three sets of IE cell lines (but not the delayed-early cell lines) responded to virus infection both in the presence of cycloheximide and with mutants defective in IE gene expression, demonstrating specific trans-activation by the pre-IE virion factor. In contrast, activation in the TK hybrid cell types required viral gene expression and the presence of a functional IE175 gene product. Up to 30-fold amplification in the copy number of the resident IFN or CAT DNA sequences also occurred within 20 h after HSV infection in IE175 hybrid cells but not in TK hybrid cells. Amplification was abolished either by treatment with phosphonacetate or by superinfection with a ts mutant unable to synthesize viral DNA, demonstrating specific HSV activation of the viral DNA replication origin (oriS) present in the IE hybrid constructs.