A new GFP-tagged line reveals unexpected Otx2 protein localization in retinal photoreceptors.
A new GFP-tagged line reveals unexpected Otx2 protein localization in retinal photoreceptors.
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DOI：
10.1186/1471-213x-7-122
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发表时间：
2007-11-02
影响因子：
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通讯作者：
Lamonerie, Thomas
中科院分区：
文献类型：
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作者：
Fossat, Nicolas;Le Greneur, Coralie;Beby, Francis;Vincent, Stephane;Godement, Pierre;Chatelain, Gilles;Lamonerie, Thomas
蛋白质表达和定位的动态监测是理解生物过程的基础。成对类同源结构域的转录因子Otx2是脊椎动物头部和大脑正常发育所必需的。最近的条件性基因敲除研究指出，这种蛋白质在发育后期和出生后的生活中有多种作用。然而，后来的表达和功能仍然很难表征，因为在任何发育阶段检测蛋白质的特异性试剂仍然缺失。我们产生了一种新的小鼠品系，在Otx2编码序列中插入GFP基因，以监测基因活性，同时保留其大部分功能。我们的研究结果表明，这条线代表了一个方便的工具来捕捉从早期胚胎阶段到成年的Otx2基因表达的动态。此外，我们可以可视化Otx 2蛋白的细胞内定位。在视网膜中，我们重新解释了蛋白质分布的前一个观点，并显示了核内蛋白质定位的进一步调节水平，这取决于细胞类型。GFP标记的Otx2小鼠系完全概括了先前已知的表达模式，并带来了检测Otx2基因活性的额外准确性和容易性。这为大脑发育的高度动态演员的实时成像开辟了道路，并且可以适应任何突变背景，以探测Otx 2和突变基因之间的遗传相互作用。
Dynamic monitoring of protein expression and localization is fundamental to the understanding of biological processes. The paired-class homeodomain-containing transcription factor Otx2 is essential for normal head and brain development in vertebrates. Recent conditional knockout studies have pointed to multiple roles of this protein during late development and post-natal life. Yet, later expression and functions remain poorly characterized as specific reagents to detect the protein at any stage of development are still missing. We generated a new mouse line harbouring an insertion of the GFP gene within the Otx2 coding sequence to monitor the gene activity while preserving most of its functions. Our results demonstrate that this line represents a convenient tool to capture the dynamics of Otx2 gene expression from early embryonic stages to adulthood. In addition, we could visualize the intracellular location of Otx2 protein. In the retina, we reinterpret the former view of protein distribution and show a further level of regulation of intranuclear protein localization, which depends on the cell type. The GFP-tagged Otx2 mouse line fully recapitulates previously known expression patterns and brings additional accuracy and easiness of detection of Otx2 gene activity. This opens up the way to live imaging of a highly dynamic actor of brain development and can be adapted to any mutant background to probe for genetic interaction between Otx2 and the mutated gene.