Transcription factor veracity: Is GBF3 responsible for ABA-regulated expression of Arabidopsis Adh?
Transcription factor veracity: Is GBF3 responsible for ABA-regulated expression of Arabidopsis Adh?
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DOI：
10.1105/tpc.8.5.847
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发表时间：
1996-05-01
期刊：
影响因子：
11.6
通讯作者：
Ferl, RJ
中科院分区：
文献类型：
--
作者：
Lu, GH;Paul, AL;Ferl, RJ
在拟南芥醇脱氢酶（Adh）启动子中，G-box可能通过脱落酸（ABA）的作用来调节其在低温和脱水条件下的表达，并在细胞培养过程中与体内核蛋白复合物结合。在本报告中，我们测试了用于鉴定DNA结合蛋白的生化方法的传统智慧，并通过使用拟南芥Adh启动子的G-box和附近的半G-box元件作为模型系统来评估它们的特异性相互作用。典型的体外实验证实了G-box因子3 （GBF3）与G-box和半G-box元件的特异性相互作用，硫酸二甲酯足迹分析证实了GBF3的体外结合特征与体内在G-box检测到的足迹特征基本匹配，因为RNA凝胶印迹数据表明GBF3本身是由ABA诱导的。我们可能已经得出结论，在细胞培养中，GBFB确实是负责与Adh G-box结合的GBF，因此负责aba调节的Adh表达。这一结论的潜在局限性暴露于以下事实：其他gbf与GBF3结合的G-box具有相同的特征，体内和体外足迹特征之间的细微差异表明，除GBF3以外的因素或除GBF3外的因素与一半G-box元件相互作用。
Assignment of particular transcription factors to specific roles in promoter elements can be problematic, especially in systems such as the G-box, where multiple factors of overlapping specificity exist, In the Arabidopsis alcohol dehydrogenase (Adh) promoter, the G-box regulates expression in response to cold and dehydration, presumably through the action of abscisic acid (ABA), and is bound by a nuclear protein complex in vivo during expression in cell cultures. In this report, we test the conventional wisdom of biochemical approaches used to identify DNA binding proteins and assess their specific interactions by using the G-box and a nearby half G-box element of the Arabidopsis Adh promoter as a model system. Typical in vitro assays demonstrated specific interaction of G-box factor 3 (GBF3) with both the G-box and the half G-box element, Dimethyl sulfate footprint analysis confirmed that the in vitro binding signature of GBF3 essentially matches the footprint signature detected in vivo at the G-box, Because RNA gel blot data indicated that GBF3 is itself induced by ABA, we might have concluded that GBFB is indeed the GBF responsible in cell cultures for binding to the Adh G-box and is therefore responsible for ABA-regulated expression of Adh. Potential limitations of this conclusion are exposed by the fact that other GBFs bind the G-box with the same signature as GBF3, and subtle differences between in vivo and in vitro footprint signatures indicate that factors other than or in addition to GBF3 interact with the half G-box element.