Real-time monitoring of functional interactions between upstream and core promoter sequences in living cells of sea urchin embryos
Real-time monitoring of functional interactions between upstream and core promoter sequences in living cells of sea urchin embryos
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DOI：
10.1093/nar/gkm519
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发表时间：
2007-07-01
影响因子：
14.9
通讯作者：
Kokubo, Tetsuro
中科院分区：
文献类型：
--
作者：
Kobayashi, Akiko;Watanabe, Youko;Kokubo, Tetsuro
在酵母、果蝇和哺乳动物细胞中，上游启动子序列和核心启动子序列在转录激活方面存在一些功能相容性。在这里，我们研究了海胆胚胎中是否存在类似的相容性，如果是这样，它们是否在早期发育过程中受到动态调节。两个报告质粒，每个含有一个测试启动子共轭CFP或YFP，同时引入到胚胎中，和他们的表达模式进行了研究，通过荧光显微镜。马粪海胆（Hemicentrotus pulcherrimus，Hp）OtxE启动子的上游序列驱动其自身的核心启动子和紫球海胆（Strongylocentrotus purpuratus，Sp）Spec2a的核心启动子在不同的胚胎区域，特别是在原肠胚晚期的表达。有趣的是，当这个上游序列的四个假定的转录因子结合位点被单独突变时，所得到的序列从同一组的两个核心启动子指导不同的时空表达，这表明上游因素的组合可以决定海胆胚胎中核心启动子的使用。此外，无论上游序列是完整的还是突变的，共有或非共有TATA序列的插入或缺失都会显著改变表达谱。因此，TATA序列可以作为这些细胞中核心启动子选择的主要决定因素。
There are some functional compatibilities between upstream and core promoter sequences for transcriptional activation in yeast, Drosophila and mammalian cells. Here we examined whether similar compatibilities exist in sea urchin embryos, and if so, whether they are dynamically regulated during early development. Two reporter plasmids, each containing a test promoter conjugated to either CFP or YFP, were concurrently introduced into embryos, and their expression patterns were studied by fluorescence microscopy. The upstream sequence of the Hemicentrotus pulcherrimus ( Hp) OtxE promoter drives the expression of its own core promoter and that of Strongylocentrotus purpuratus (Sp) Spec2a in different embryonic regions, especially at the late gastrula stage. Interestingly, when the four putative transcription factor binding sites of this upstream sequence were individually mutated, the resulting sequences directed different spatio-temporal expression from the same set of two core promoters, indicating that combinations of upstream factors may determine core promoter usage in sea urchin embryos. In addition, the insertion or deletion of consensus or nonconsensus TATA sequences changed the expression profile significantly, irrespective of whether the upstream sequence was intact or mutated. Thus, the TATA sequence may serve as a primary determinant for core promoter selection in these cells.