Yan Hua, Chretien Robert, Ye Jingsong, Rommens Caius M
Simplot Plant Sciences, J.R. Simplot Company, Boise, Idaho 83706, USA.
Plant Physiol. 2006 Aug;141(4):1508-18. doi: 10.1104/pp.106.082271. Epub 2006 Jun 9.
An important component of conventional sense, antisense, and double-strand RNA-based gene silencing constructs is the transcriptional terminator. Here, we show that this regulatory element becomes obsolete when gene fragments are positioned between two oppositely oriented and functionally active promoters. The resulting convergent transcription triggers gene silencing that is at least as effective as unidirectional promoter-to-terminator transcription. In addition to short, variably sized, and nonpolyadenylated RNAs, terminator-free cassette produced rare, longer transcripts that reach into the flanking promoter. These read-through products did not influence the efficacy and expression levels of the neighboring hygromycin phosphotransferase gene. Replacement of gene fragments by promoter-derived sequences further increased the extent of gene silencing. This finding indicates that genomic DNA may be a more efficient target for gene silencing than gene transcripts.
传统意义上基于正义、反义及双链RNA的基因沉默构建体的一个重要组成部分是转录终止子。在此，我们表明，当基因片段位于两个方向相反且功能活跃的启动子之间时，这个调控元件就过时了。由此产生的汇聚转录引发基因沉默，其效果至少与从启动子到终止子的单向转录一样有效。除了短的、大小可变的和非聚腺苷酸化的RNA外，无终止子盒产生了罕见的、更长的转录本，这些转录本延伸到侧翼启动子。这些通读产物不影响相邻潮霉素磷酸转移酶基因的功效和表达水平。用启动子衍生序列替代基因片段进一步提高了基因沉默的程度。这一发现表明，基因组DNA可能比基因转录本更有效地成为基因沉默的靶点。
