Adoption of the Q transcriptional regulatory system for zebrafish transgenesis.
Adoption of the Q transcriptional regulatory system for zebrafish transgenesis.
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DOI：
10.1016/j.ymeth.2013.06.012
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发表时间：
2014-04-01
期刊：
影响因子：
4.8
通讯作者：
Halpern, Marnie E.
中科院分区：
文献类型：
--
作者：
Subedi, Abhignya;Macurak, Michelle;Gee, Stephen T.;Monge, Estela;Goll, Mary G.;Potter, Christopher J.;Parsons, Michael J.;Halpern, Marnie E.
酵母的 Gal4-UAS 调控系统广泛用于调节果蝇的基因表达；然而，由于富含 CpG 的多拷贝上游激活序列的逐步甲基化和沉默，其在转基因斑马鱼中的用途受到限制。尽管经过修饰的、重复性较低的 UAS 构建体可以克服这个问题，但非常需要有额外的转录调控系统，可以独立应用或与 Gal4/UAS 系统组合应用以进行交叉基因表达。粗糙脉孢菌的 Q 转录调控系统的功能与 Gal4/UAS 类似。 QF 是一种转录激活剂，可与 QUAS 上游调控序列结合以驱动报告基因表达。与 Gal4 不同，QF 结合位点不包含易受 DNA 甲基化影响的必需 CpG 二核苷酸序列。 QS 蛋白是 QF 介导的转录激活的阻遏蛋白，类似于 Gal80。 Q 系统的功能已在果蝇和线虫中得到证明，我们现在报告其在脊椎动物模型斑马鱼、斑马鱼中的成功应用。几个组织特异性启动子被用来驱动稳定转基因系中的 QF 表达，通过 QUAS:GFP 转基因的激活进行评估。研究发现，QS 阻遏蛋白可显着降低注射斑马鱼胚胎中的 QF 活性；然而，在普遍存在的启动子控制下表达QS的转基因动物中尚未实现类似的抑制。还生成了包含 QUAS 和 UAS 的双报告构建体，每个上游都有不同的荧光蛋白，并在瞬时测定中进行测试，证明这两个系统可以在同一细胞内并行工作。 Q系统的采用应该会大大增加转基因方法调节斑马鱼基因表达的多功能性和力量。
The Gal4-UAS regulatory system of yeast is widely used to modulate gene expression in Drosophila; however, there are limitations to its usefulness in transgenic zebrafish, owing to progressive methylation and silencing of the CpG-rich multicopy upstream activation sequence. Although a modified, less repetitive UAS construct may overcome this problem, it is highly desirable to have additional transcriptional regulatory systems that can be applied independently or in combination with the Gal4/UAS system for intersectional gene expression. The Q transcriptional regulatory system of Neurospora crassa functions similarly to Gal4/UAS. QF is a transcriptional activator that binds to the QUAS upstream regulatory sequence to drive reporter gene expression. Unlike Gal4, the QF binding site does not contain essential CpG dinucleotide sequences that are subject to DNA methylation. The QS protein is a repressor of QF mediated transcriptional activation akin to Gal80. The functionality of the Q system has been demonstrated in Drosophila and C. elegans and we now report its successful application to a vertebrate model, the zebrafish, Danio rerio. Several tissue-specific promoters were used to drive QF expression in stable transgenic lines, as assessed by activation of a QUAS:GFP transgene. The QS repressor was found to dramatically reduce QF activity in injected zebrafish embryos; however, a similar repression has not yet been achieved in transgenic animals expressing QS under the control of ubiquitous promoters. A dual reporter construct containing both QUAS and UAS, each upstream of different fluorescent proteins was also generated and tested in transient assays, demonstrating that the two systems can work in parallel within the same cell. The adoption of the Q system should greatly increase the versatility and power of transgenic approaches for regulating gene expression in zebrafish.
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影响因子：
2.6
作者：
Parsons, Michael J.;Pisharath, Harshan;Yusuff, Shamila;Moore, John C.;Siekmann, Arndt F.;Lawson, Nathan;Leach, Steven D.
通讯作者：
Leach, Steven D.
影响因子：
64.5
作者：
Potter CJ;Tasic B;Russler EV;Liang L;Luo L
通讯作者：
Luo L
影响因子：
2.7
作者：
Akitake, Courtney M.;Macurak, Michelle;Goll, Mary G.
通讯作者：
Goll, Mary G.
DOI：
10.1073/pnas.97.21.11403
发表时间：
2000-10-10
影响因子：
11.1
作者：
Kawakami, K;Shima, A;Kawakami, N
通讯作者：
Kawakami, N
影响因子：
2.7
作者：
AMSTERDAM, A;LIN, S;HOPKINS, N
通讯作者：
HOPKINS, N