BACULOVIRUS GP64 GENE-EXPRESSION - ANALYSIS OF SEQUENCES MODULATING EARLY TRANSCRIPTION AND TRANSACTIVATION BY IE1
BACULOVIRUS GP64 GENE-EXPRESSION - ANALYSIS OF SEQUENCES MODULATING EARLY TRANSCRIPTION AND TRANSACTIVATION BY IE1
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DOI：
10.1128/jvi.65.11.5820-5827.1991
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发表时间：
1991-11-01
影响因子：
5.4
通讯作者：
ROHRMANN, GF
中科院分区：
文献类型：
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作者：
BLISSARD, GW;ROHRMANN, GF
杆状病毒主要包膜糖蛋白基因（gp 64）的表达受早期和晚期启动子的转录调控。为了表征早期启动子和识别参与调节gp 64早期转录的序列，从Orygia pseudotsugata核型多角体病毒gp 64启动子产生启动子-报告基因融合，并通过在未感染的昆虫细胞中瞬时表达进行分析。对于这些分析，在gp 64上游调控区中构建5'缺失突变。较大的启动子结构在未感染的舞毒蛾细胞中是有功能的，表明从gp 64早期启动子的转录不需要额外的病毒基因产物。gp 64上游区域的缺失分析揭示了几个调控区域。这些包括在-319和-166个核苷酸（nt）之间的推定负调控元件和在-166和-77 nt之间的多个正调控元件。位于-77和-62 nt之间的TATA盒的缺失导致转录活性的丧失。共转染的报告构建体和含有杆状病毒转录反式激活因子基因（苜蓿银纹夜蛾核型多角体病毒IE 1）的质粒导致转录反式激活的所有结构含有完整的TATA框。这些数据表明，序列上游的gp 64 TATA框是不是必不可少的IE 1的反式激活和早期转录起始位点上游只有34个核苷酸是必要的基础水平的转录和由IE 1的反式激活。还在草地贪夜蛾和黑腹果蝇的细胞系中检查了gp 64早期启动子的功能。
Expression of the baculovirus major envelope glycoprotein gene (gp64) is regulated by transcription from both early and late promoters. To characterize the early promoter and identify sequences involved in the regulation of gp64 early transcription, promoter-reporter gene fusions were generated from the Orygia pseudotsugata nuclea polyhedrosis virus gp64 promoter and were analyzed by transient expression in uninfected insect cells. For these analyses, 5' deletion mutations were constructed in the gp64 upstream regulatory region. Larger promoter constructs were functional in uninfected Lymantria dispar cells, indicating that transcription from the gp64 early promoter required no additional viral gene products. Deletion analysis of the gp64 upstream region revealed several regulatory regions. These included a putative negative regulatory element between -319 and -166 nucleotides (nt) and multiple positive regulatory elements between -166 and -77 nt. Deletion of the TATA box located between -77 and -62 nt resulted in the loss of transcriptional activity. Cotransfections of reporter constructs and a plasmid containing a baculovirus transcriptional transactivator gene (Autographa californica nuclear polyhedrosis virus IE1) resulted in transcriptional transactivation of all constructs containing an intact TATA box. These data demonstrate that sequences upstream of the gp64 TATA box are not essential for IE1 transactivation and that only 34 nt upstream of the early transcription start site were necessary for basal levels of transcription and for transactivation by IE1. Function of the gp64 early promoter was also examined in cell lines from Spodoptera frugiperda and Drosophila melanogaster.