Palla F, Casano C, Albanese I, Anello L, Gianguzza F, Di Bernardo M G, Bonura C, Spinelli G
Dipartimento di Biologia Cellulare e dello Sviluppo, Palermo, Italy.
Proc Natl Acad Sci U S A. 1989 Aug;86(16):6033-7. doi: 10.1073/pnas.86.16.6033.
Functional tests, performed by microinjection into Xenopus laevis oocytes, show that a DNA fragment containing the modulator of the early histone H2A gene of Paracentrotus lividus enhances transcription of a reporter gene when located, in the physiological orientation, upstream of the tk basal promoter. Gel retardation and DNase I footprinting assays further reveal that the H2A modulator contains at least two binding sites [upstream sequence elements 1 and 2 (USE 1 and USE 2)] for nuclear factors extracted from sea urchin embryos, which actively transcribe the early histone gene set. Interestingly, USE 1 is highly homologous to a cis-acting element previously identified in the H2A modulator of Psammechinus miliaris [Grosschedl, R., Mächler, M., Rohrer, U. & Birnstiel, M. L. (1983) Nucleic Acids Res. 11, 8123-8136]. Finally, a cloned oligonucleotide containing the USE 1 sequence competes efficiently in Xenopus oocytes with the H2A modulator to prevent enhancement of transcription of the reporter gene. From these results, we conclude that USE 1 and perhaps USE 2 in the H2A modulator are upstream transcriptional elements that are recognized by trans-acting factors common to Xenopus and sea urchin.
通过显微注射到非洲爪蟾卵母细胞中进行的功能测试表明，一个包含紫球海胆早期组蛋白H2A基因调节子的DNA片段，当以生理方向位于tk基础启动子上游时，可增强报告基因的转录。凝胶阻滞和DNase I足迹分析进一步揭示，H2A调节子包含至少两个与从海胆胚胎中提取的核因子的结合位点[上游序列元件1和2（USE 1和USE 2）]，这些核因子可积极转录早期组蛋白基因集。有趣的是，USE 1与先前在细雕刻肋海胆的H2A调节子中鉴定出的顺式作用元件高度同源[格罗斯切尔德，R.，马赫勒，M.，罗勒，U.和伯恩施泰尔，M. L.（1983年）《核酸研究》11，8123 - 8136]。最后，一个包含USE 1序列的克隆寡核苷酸在非洲爪蟾卵母细胞中与H2A调节子有效竞争，以阻止报告基因转录的增强。从这些结果中，我们得出结论，H2A调节子中的USE 1以及可能的USE 2是上游转录元件，可被非洲爪蟾和海胆共有的反式作用因子识别。
