Mosca J D, Jeang K T, Pitha P M, Hayward G S
J Virol. 1987 Mar;61(3):819-28. doi: 10.1128/JVI.61.3.819-828.1987.
We have constructed stable DNA-transfected LTK+ cell lines containing two different coselected hybrid interferon (IFN) genes driven by the usually strong and constitutive promoter from the immediate-early 94K protein (IE94) gene of simian cytomegalovirus. Surprisingly, and unlike hybrid IE94-chloramphenicol acetyltransferase gene constructs, both of the IE94-IFN genes (one with and one without the complex spliced intron region) produced relatively low basal titers of biologically active human IFN in the mouse cell lines. However, IFN expression could be stimulated up to 120-fold by superinfection with herpes simplex virus (HSV), although not with cytomegalovirus. To examine the mechanism of this unexpected HSV induction process, we measured the levels of both IE94-IFN mRNA and IFN protein produced under various infection protocols. Compared with similar previously characterized cell lines containing hybrid IFN genes under the control of HSV IE or delayed-early (DE) promoters, activation of IFN expression first occurred at an intermediate time. Both IE94-IFN cell lines also produced an unusual pattern of response to infection with the HSV IE regulation-deficient mutants tsK and tsB7: stimulation of IFN synthesis occurred in the absence of a functional HSV IE175 (or ICP4) gene product, but did not occur in the absence of uncoating of virus capsids. Cycloheximide treatment (without virus infection) also gave a rapid 30-fold increase in steady-state levels of correctly initiated mRNA from both types of IE94-IFN hybrid genes, but had no effect on cells containing the IE175-IFN construct. Therefore, we conclude that the use of the IE94-IFN constructs identifies a novel HSV regulatory response that requires a previously unrecognized function of HSV and does not involve either IE175 or the pre-IE "virion factor" trans-activators that are known to stimulate transcription of HSV IE and DE genes, respectively.
我们构建了稳定的DNA转染的LTK⁺细胞系，这些细胞系包含两个不同的共选择杂交干扰素（IFN）基因，由来自猿猴巨细胞病毒立即早期94K蛋白（IE94）基因的通常很强的组成型启动子驱动。令人惊讶的是，与杂交IE94 - 氯霉素乙酰转移酶基因构建体不同，两个IE94 - IFN基因（一个有复杂剪接内含子区域，一个没有）在小鼠细胞系中产生的具有生物活性的人IFN基础滴度相对较低。然而，单纯疱疹病毒（HSV）的超感染可将IFN表达刺激高达120倍，而巨细胞病毒则不能。为了研究这种意外的HSV诱导过程的机制，我们测量了在各种感染方案下产生的IE94 - IFN mRNA和IFN蛋白的水平。与先前表征的在HSV IE或延迟早期（DE）启动子控制下包含杂交IFN基因的类似细胞系相比，IFN表达的激活首先发生在中间时间。两个IE94 - IFN细胞系对HSV IE调节缺陷突变体tsK和tsB7的感染也产生了不寻常的反应模式：在没有功能性HSV IE175（或ICP4）基因产物的情况下发生IFN合成的刺激，但在没有病毒衣壳脱壳的情况下不发生。环己酰亚胺处理（无病毒感染）也使两种类型的IE94 - IFN杂交基因正确起始的mRNA的稳态水平迅速增加30倍，但对含有IE175 - IFN构建体的细胞没有影响。因此，我们得出结论，使用IE94 - IFN构建体鉴定出一种新的HSV调节反应，该反应需要HSV以前未被认识的功能，并且不涉及分别已知刺激HSV IE和DE基因转录的IE175或前IE“病毒体因子”反式激活因子。
