Identification of an evolutionarily conserved regulatory element of the zebrafish col2a1a gene.
Identification of an evolutionarily conserved regulatory element of the zebrafish col2a1a gene.
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斑马鱼COL2A1A基因的进化保守调节元件的鉴定。
DOI：
10.1016/j.ydbio.2011.06.020
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发表时间：
2011-09-15
影响因子：
2.7
通讯作者：
Topczewski J
中科院分区：
文献类型：
--
作者：
Dale RM;Topczewski J
斑马鱼（Danio rerio）是研究脊椎动物发育（包括骨骼发生）的优良模式生物。通过使用胶原II型α 1 （Col2a1）基因的软骨特异性调节元件，哺乳动物软骨形成的研究取得了很大进展。为了在斑马鱼中分离出这样一个元件，我们比较了两种col2a1同源物的表达，发现col2a1b（一种以前未被表征的斑马鱼同源物）的表达仅与col2a1a部分重叠。我们的分析重点是col2a1a，因为它在堆叠的软骨细胞和软骨膜中都有表达。通过比较几种硬骨鱼col2a1a预测转录起始位点周围的基因组序列，我们确定了一个位于推定转录起始位点上游1.7 kb的小高度保守序列（R2）。有趣的是，该元件的序列和位置在硬骨鱼和哺乳动物Col2a1之间都不保守。我们只使用R2元件或转录起始位点的整个1.7 kb片段5 '，获得了瞬时和稳定的转基因系。鉴定出的调控元件能够通过驱动颅面软骨、耳、脊索、底板、下索和鳍中报告基因的强劲表达，以类似于内源性col2a1a的表达模式，跟踪各种组织中的细胞发育。利用R2调控元件驱动的报告基因，我们分析了脊索鞘细胞从最初均匀的细胞堆叠中退出并包裹膨胀的空泡状脊索细胞时的形态发生。最后，我们发现像内源性col2a1a一样，这些报告基因的颅面表达依赖于Sox9a转录因子的活性。同时，Sox9a敲低后脊索表达得以维持，提示其他因子可以通过该组织中所鉴定的调控元件激活表达。
Zebrafish (Danio rerio) is an excellent model organism for the study of vertebrate development including skeletogenesis. Studies of mammalian cartilage formation were greatly advanced through the use of a cartilage specific regulatory element of the Collagen type II alpha 1 (Col2a1) gene. In an effort to isolate such an element in zebrafish, we compared the expression of two col2a1 homologues and found that expression of col2a1b, a previously uncharacterized zebrafish homologue, only partially overlaps with col2a1a. We focused our analysis on col2a1a, as it is expressed in both the stacked chondrocytes and the perichondrium. By comparing the genomic sequence surrounding the predicted transcriptional start site of col2a1a among several species of teleosts we identified a small highly conserved sequence (R2) located 1.7 kb upstream of the presumptive transcriptional initiation site. Interestingly, neither the sequence nor location of this element is conserved between teleost and mammalian Col2a1. We generated transient and stable transgenic lines with just the R2 element or the entire 1.7 kb fragment 5’ of the transcriptional initiation site. The identified regulatory elements enable the tracking of cellular development in various tissues by driving robust reporter expression in craniofacial cartilage, ear, notochord, floor plate, hypochord and fins in a pattern similar to the expression of endogenous col2a1a. Using a reporter gene driven by the R2 regulatory element, we analyzed the morphogenesis of the notochord sheath cells as they withdraw from the stack of initially uniform cells and encase the inflating vacuolated notochord cells. Finally, we show that like endogenous col2a1a, craniofacial expression of these reporter constructs depends on Sox9a transcription factor activity. At the same time, notochord expression is maintained after Sox9a knockdown, suggesting that other factors can activate expression through the identified regulatory element in this tissue.
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影响因子：
10.5
作者：
Akiyama, H;Chaboissier, MC;de Crombrugghe, B
通讯作者：
de Crombrugghe, B
影响因子：
5.3
作者：
Han, Yu;Lefebvre, Veronique
通讯作者：
Lefebvre, Veronique
DOI：
10.1111/j.1096-3642.2003.00097.x
发表时间：
2004-02-01
影响因子：
2.8
作者：
Grande, T;Young, B
通讯作者：
Young, B
影响因子：
2.7
作者：
Kague, Erika;Bessling, Seneca L.;Fisher, Shannon
通讯作者：
Fisher, Shannon
影响因子：
14.9
作者：
CORPET, F
通讯作者：
CORPET, F