Bandyopadhyay R S, Bruce W B, Gurley W B
Department of Microbiology and Cell Science, University of Florida, Gainsville 32611.
J Biol Chem. 1989 Nov 15;264(32):19399-406.
DNA sequences involved in the expression of the agropine synthase gene (ags) of T-DNA were identified by analysis of transcriptional activity of promoter mutants in crown gall tumors of sunflower. Precise quantification of activity was achieved using a homologous reference gene as an internal standard. Analysis of 5'-deletion mutants demonstrated the requirement of 314 base pairs of upstream DNA sequences for optimal activity. Five regions involved in transcriptional regulation were identified in the 5'-flanking sequences between positions -74 and -314. Four of these regions make a positive contribution to promoter activity, and the fifth exerts a negative influence. The TATA motif (-26 to -33) and the TATA proximal domain (-74 to -105), which contains two sequences similar to the mammalian CCAAT box, are the major determinants of promoter activity. The two TATA distal domains A and B are separated by a negative element (-166 to -205) which may attenuate promoter strength by distancing the TATA distal domain B (-206 to -314 base pairs) from downstream components of the promoter. The TATA distal domain B contains the a/b repeat first described in the nopaline synthase (nos) promoter and was unable to support transcription in the absence of elements within the TATA proximal domain.
通过对向日葵冠瘿瘤中启动子突变体的转录活性进行分析，确定了参与T-DNA农杆碱合酶基因（ags）表达的DNA序列。使用同源参考基因作为内标实现了活性的精确量化。对5'-缺失突变体的分析表明，最佳活性需要314个碱基对的上游DNA序列。在-74至-314位之间的5'-侧翼序列中鉴定出五个参与转录调控的区域。其中四个区域对启动子活性有正向作用，第五个区域则起负向影响。TATA基序（-26至-33）和TATA近端结构域（-74至-105）包含两个与哺乳动物CCAAT框相似的序列，是启动子活性的主要决定因素。两个TATA远端结构域A和B被一个负向元件（-166至-205）隔开，该负向元件可能通过使TATA远端结构域B（-206至-314碱基对）与启动子的下游元件保持距离来减弱启动子强度。TATA远端结构域B包含首次在胭脂碱合酶（nos）启动子中描述的a/b重复序列，并且在没有TATA近端结构域内元件的情况下无法支持转录。
