DeLuca N A, Courtney M A, Schaffer P A
J Virol. 1984 Dec;52(3):767-76. doi: 10.1128/JVI.52.3.767-776.1984.
A large number of temperature-sensitive (ts) mutants of herpes simplex virus type 1 (HSV-1) in the gene encoding the immediate-early transcriptional regulatory protein, ICP4, have been isolated and characterized with respect to expression of the immediate-early, early, and late viral gene products. The hallmark of these mutants is the overproduction of immediate-early gene products and the underproduction of early and late gene products. The present study involves the preliminary genetic and molecular characterization of two unique regulatory mutants of HSV-1, ts48 and ts303. Genetically, both mutants exhibit inefficient complementation with eight ts mutants in complementation group 1-2, which defines the gene for ICP4, and marker rescue experiments place the mutations in both mutants in the 3' portion of the coding sequence for ICP4. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of ts48- and ts303-infected cell polypeptides synthesized at the nonpermissive temperature demonstrates that immediate-early polypeptides ICP4 and ICP27 are overproduced, with the simultaneous production of early polypeptides ICP6, ICP8, gB, and others. Immediate-early polypeptides are resynthesized upon temperature shift-up early in infection; however, shift-up late in infection does not result in the resynthesis of immediate-early polypeptides. Late gene products are either absent or underrepresented under long-term labeling conditions. To examine the effects of the mutations in ts48, ts303, and other ICP4 mutants specifically on early gene expression, trans-induction experiments were performed in cells transfected with the gene for chloramphenicol acetyltransferase under early gene control (tk) and superinfected with KOS, tsB32, ts48, and ts303. Mutant tsB32 did not induce chloramphenicol acetyltransferase activity above the basal level; however, ts48 and ts303 induced chloramphenicol acetyltransferase activity nearly equal to wild-type levels. Fifteen to fifty percent of wild-type levels of viral DNA are synthesized at the nonpermissive temperature in ts48- and ts303-infected cells, indicating that immediate-early and early gene functions are intact (or nearly so) and that the block in ts48 and ts303 is in a regulatory event subsequent to that exhibited by other mutants in complementation group 1-2 which are DNA-.
已分离出大量单纯疱疹病毒1型（HSV-1）的温度敏感（ts）突变体，这些突变体在编码立即早期转录调节蛋白ICP4的基因中，并且已针对立即早期、早期和晚期病毒基因产物的表达进行了表征。这些突变体的标志是立即早期基因产物过量产生，而早期和晚期基因产物产生不足。本研究涉及HSV-1的两个独特调节突变体ts48和ts303的初步遗传和分子表征。从遗传学角度来看，这两个突变体与互补组1-2中的八个ts突变体表现出低效互补，互补组1-2定义了ICP4的基因，并且标记拯救实验将这两个突变体中的突变定位在ICP4编码序列的3'部分。对在非允许温度下合成的ts48和ts303感染细胞多肽进行十二烷基硫酸钠-聚丙烯酰胺凝胶电泳分析表明，立即早期多肽ICP4和ICP27过量产生，同时产生早期多肽ICP6、ICP8、gB等。在感染早期温度升高时，立即早期多肽会重新合成；然而，在感染后期温度升高不会导致立即早期多肽的重新合成。在长期标记条件下，晚期基因产物要么不存在，要么表达不足。为了具体研究ts48、ts303和其他ICP4突变体中的突变对早期基因表达的影响，在用早期基因控制（tk）下的氯霉素乙酰转移酶基因转染并被KOS、tsB32、ts48和ts303超感染的细胞中进行了反式诱导实验。突变体tsB32没有诱导氯霉素乙酰转移酶活性超过基础水平；然而，ts48和ts303诱导的氯霉素乙酰转移酶活性几乎与野生型水平相当。在ts48和ts303感染的细胞中，在非允许温度下合成的病毒DNA水平为野生型水平的15%至50%，这表明立即早期和早期基因功能是完整的（或几乎完整），并且ts48和ts303中的阻断发生在互补组1-2中其他DNA - 突变体所表现出的调节事件之后。
