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首页> 外文期刊>Molecular and Cellular Biology >Identification In Vivo of Different Rate-Limiting Steps Associated with Transcriptional Activators in the Presence and Absence of a GAGA Element
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Identification In Vivo of Different Rate-Limiting Steps Associated with Transcriptional Activators in the Presence and Absence of a GAGA Element

机译：在存在和不存在GAGA元素的情况下与转录激活剂相关的不同速率限制步骤的体内鉴定

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We analyzed the impact of a GAGA element on a transgenic promoter in Drosophila melanogaster that was activated by proteins composed of the Teton DNA binding domain and either the heat shock factor (HSF) activation domain or a potent subdomain of VP16. Permanganate footprinting was used to monitor polymerase II (Pol II) on the transgenic promoters in vivo. Activation by Teton-HSF but not by Teton-VP16A2 required the GAGA element; this correlated with the ability of the GAGA element to establish a paused Pol II. Although the GAGA element was not required for activation by Teton-VP16A2, the GAGA element greatly accelerated the rate of activation. The permanganate data also provided evidence that Pol II encountered different rate-limiting steps, following initiation in the presence of Teton-HSF and Teton-VP16A2. The rate-limiting step in the presence of Teton-HSF was release of Pol II paused about 20 to 40 nucleotides downstream from the start site. The rate-limiting step in the presence of Teton-VP16A2 occurred much closer to the transcription start site. Several biochemical studies have provided evidence for a structural transition shortly after Pol II initiates transcription. The behavior of Pol II in the presence of Teton-VP16A2 provides the first evidence that this transition occurs in vivo.
机译：我们分析了GAGA元素对在果蝇 melanogaster 的转基因启动子上的影响，该Melanogaster 由由Tet 上的TET 上的蛋白质和DNA结合结构域组成的蛋白质激活热休克因子（HSF）激活域或VP16的有效子域。高锰酸盐的脚印用于监测体内转基因启动子的聚合酶II（POL II）。通过TET 在 -hsf上激活，但不是tet 上的 -vp16 a2 所需gaga元素;这种情况与Gaga元素建立暂停的pol II的能力相关。虽然TET 在 -vp16 a2 上不需要激活Gaga元素，但Gaga元素大大加速了激活速率。 Manganate数据还提供了证据，即POL II在TET 在 -HSF和TET 上的存在下启动后遇到不同的速率限制步骤 -vp16 a2 < / sub>。 TET 在 -HSF上存在的速率限制步骤在起始位点下暂停的POL II的释放持续约20至40个核苷酸。在 -vp16 a2 上存在的速率限制步骤 -vp16 a2 更接近转录开始站点。在POL II启动转录后，几项生化研究已经提供了结构过渡的证据。 POL II在TET 上的存在的行为 -Vp16 a2 提供了第一种发出这种转变在体内发生的证据。

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