Inhibition of endogenous MTF-1 signaling in zebrafish embryos identifies novel roles for MTF-1 in development.
Inhibition of endogenous MTF-1 signaling in zebrafish embryos identifies novel roles for MTF-1 in development.
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斑马鱼胚胎中内源性 MTF-1 信号传导的抑制确定了 MTF-1 在发育中的新作用。
DOI：
10.1016/j.bbamcr.2014.04.015
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发表时间：
2014
期刊：
影响因子：
--
通讯作者：
Jenny,MatthewJ
中科院分区：
文献类型：
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作者：
O'Shields,Britton;McArthur,AndrewG;Holowiecki,Andrew;Kamper,Martin;Tapley,Jeffrey;Jenny,MatthewJ
金属响应元件结合转录因子-1 （MTF-1）响应锌暴露或重金属或氧相关应激引起的内源性锌稳态破坏引起的细胞锌水平变化。在这里，我们报告了完整的斑马鱼MTF-1的功能特征，并与先前鉴定的缺乏高度保守的富含半胱氨酸的基序（Cys-X-Cys-Cys-X-Cys）的异构体进行了比较，这些异构体在所有其他脊椎动物MTF-1同源物中发现。为了开发新的分子工具，我们产生了一个组成核显性阴性MTF-1 （dnMTF-1）作为抑制内源性MTF-1信号的工具。通过将体外转录的dnMTF-1 mRNA微注射到斑马鱼胚胎（1-2细胞期），然后使用Agilent 4x44K阵列在28-和36-hpf胚胎上进行转录组学分析，确定dnMTF-1的体内功效。共有594个和560个探针分别在28 hpf和36 hpf时被鉴定为差异表达，在时间点之间存在有趣的重叠。受MTF-1信号抑制影响的基因主要有：核受体和参与应激信号、神经发生、肌肉发育和收缩、眼睛发育和金属稳态的基因，包括铁和血红素稳态的新发现。最后，我们研究了MTF-1在斑马鱼早期发育过程中转录组学分析鉴定的潜在新靶基因中的转录激活因子和转录抑制因子作用。
The metal responsive element-binding transcription factor-1 (MTF-1) responds to changes in cellular zinc levels caused by zinc exposure or disruption of endogenous zinc homeostasis by heavy metals or oxygen-related stress. Here we report the functional characterization of a complete zebrafish MTF-1 in comparison with the previously identified isoform lacking the highly conserved cysteine-rich motif (Cys-X-Cys-Cys-X-Cys) found in all other vertebrate MTF-1 orthologs. In an effort to develop novel molecular tools, a constitutively nuclear dominant-negative MTF-1 (dnMTF-1) was generated as tool for inhibiting endogenous MTF-1 signaling. Thein vivoefficacy of the dnMTF-1 was determined by microinjectingin vitrotranscribed dnMTF-1 mRNA into zebrafish embryos (1–2 cell stage) followed by transcriptomic profiling using an Agilent 4x44K array on 28- and 36-hpf embryos. A total of 594 and 560 probes were identified as differentially expressed at 28 hpf and 36 hpf, respectively, with interesting overlaps between timepoints. The main categories of genes affected by the inhibition of MTF-1 signaling were: nuclear receptors and genes involved in stress signaling, neurogenesis, muscle development and contraction, eye development, and metal homeostasis, including novel observations in iron and heme homeostasis. Finally, we investigate both the transcriptional activator and transcriptional repressor role of MTF-1 in potential novel target genes identified by transcriptomic profiling during early zebrafish development.