Gutiérrez Serafín, Kikhno Iryna, López Ferber Miguel
Laboratoire de Pathologie Comparée, UR 1231 INRA, 30380 Saint Christol les Alès, France.
J Gen Virol. 2004 Feb;85(Pt 2):331-341. doi: 10.1099/vir.0.19623-0.
The pif gene (per os infectivity factor) of Spodoptera littoralis nucleopolyhedrovirus (SpliNPV) encodes a structural protein essential for oral infection. This protein is expressed in very low quantities. In this study, transcription and promoter analysis of SpliNPV pif were carried out to understand more fully the regulation of pif gene expression. Transcription in the pif gene region was examined using RT-PCR, Northern blot, primer extension, ribonuclease protection and 3' RACE. The pif gene was encoded by a late bicistronic messenger, which was characterized. This 1.9 kb messenger was present in very small amounts. In addition, this messenger was part of a set of six late mRNAs overlapping the pif sequence. A functional complementation assay was used to analyse the pif promoter. This assay allowed the detection of amounts of PIF which were sufficient for the production of orally infectious virions. The 13 bp region upstream from the initial ATG of pif was required and sufficient for the production of orally infectious virions. This promoter region was much shorter than the studied baculovirus promoters. A late promoter motif (TTAAG) is situated at the 5' end of this region. This motif was shown to be the promoter core by using single mutations of the motif in the complementation assay. These results suggest that the low expression of the pif gene is regulated chiefly at the transcriptional level.
埃及棉铃虫核型多角体病毒（SpliNPV）的pif基因（经口感染性因子）编码一种对经口感染至关重要的结构蛋白。这种蛋白的表达量非常低。在本研究中，对SpliNPV pif进行了转录和启动子分析，以更全面地了解pif基因表达的调控。使用逆转录聚合酶链反应（RT-PCR）、Northern印迹、引物延伸、核糖核酸酶保护和3' 末端快速扩增（3' RACE）检测pif基因区域的转录情况。pif基因由一个晚期双顺反子信使RNA编码，并对其进行了表征。这种1.9 kb的信使RNA含量极少。此外，该信使RNA是与pif序列重叠的一组六个晚期mRNA的一部分。使用功能互补试验分析pif启动子。该试验能够检测到足以产生经口感染性病毒粒子的PIF量。pif起始ATG上游13 bp的区域对于产生经口感染性病毒粒子是必需且足够的。该启动子区域比所研究的杆状病毒启动子短得多。一个晚期启动子基序（TTAAG）位于该区域的5' 端。通过在互补试验中对该基序进行单突变，表明该基序是启动子核心。这些结果表明，pif基因的低表达主要在转录水平受到调控。
