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首页> 外文期刊>Protoplasma: An International Journal of Cell Biology >The DnaJ-like zinc finger domain protein ORANGE localizes to the nucleus in etiolated cotyledons of Arabidopsis thaliana
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The DnaJ-like zinc finger domain protein ORANGE localizes to the nucleus in etiolated cotyledons of Arabidopsis thaliana

机译：DnaJ样锌指结构域蛋白ORANGE定位于拟南芥黄化子叶的核中。

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摘要

Vitamin A deficiency (VAD) is a worldwide health problem. Overexpression of the DnaJ-like zinc finger domain protein ORANGE (OR) is a novel strategy for the biofortification of pro-vitamin A carotenoids in different staple crops to alleviate VAD. In plants, OR triggers the differentiation from non-pigmented plastids into carotenoid-accumulating plastids. There are different reports on the subcellular localization of this protein in either chloroplasts or the nucleus, both of which were supported by confocal observation and protein-protein interaction results. In this work, we studied the subcellular localization of OR in the cotyledons of germinating seedlings whose plastids were transitioning from non-pigmented proplastids into carotenoid-accumulating etioplasts in the dark, and then into chloroplasts upon illumination. Our Western blot analysis identified two bands of the Arabidopsis OR protein (AtOR) from the chloroplast fraction of the mature leaves (i.e., a 34-kDa form corresponding to the full-length peptide and a 30-kDa form suggesting the removal of the N-terminal chloroplast transit peptide). We found that the full-length AtOR was predominantly localized in the nucleus in etiolated cotyledons, although its abundance decreased upon illumination. Our bioinformatics analysis indicated a nuclear localization signal (NLS) after the N-terminal chloroplast transit peptide. When we substituted different N-terminal regions of AtOR with the green fluorescent protein, our confocal observations demonstrated that this NLS was sufficient to target AtOR to the nucleus. Our results demonstrate that AtOR is a dual-targeted protein that mainly localizes in the nucleus in etiolated cotyledons.
机译：维生素A缺乏症（VAD）是全球性的健康问题。 DnaJ样锌指结构域蛋白ORANGE（OR）的过表达是一种生物强化维生素A类胡萝卜素在不同主食作物中缓解VAD的新策略。在植物中，OR触发从无色素质体向类胡萝卜素蓄积质体的分化。关于该蛋白在叶绿体或细胞核中的亚细胞定位的报道不同，这两者都得到了共聚焦观察和蛋白-蛋白相互作用结果的支持。在这项工作中，我们研究了发芽幼苗的子叶中OR的亚细胞定位，这些子叶的质体从无色素的质体转变为在黑暗中积累类胡萝卜素的质体，然后在光照下转化为叶绿体。我们的蛋白质印迹分析从成熟叶片的叶绿体级分中鉴定出两条拟南芥OR蛋白（AtOR）带（即对应于全长肽的34 kDa形式和30 kDa形式，表明去除了N -末端叶绿体转运肽）。我们发现全长AtOR主要定位在黄化子叶的细胞核中，尽管光照后其丰度降低了。我们的生物信息学分析表明，在N末端叶绿体转运肽之后出现了核定位信号（NLS）。当我们用绿色荧光蛋白取代AtOR的不同N末端区域时，我们的共聚焦观察表明，该NLS足以将AtOR靶向细胞核。我们的结果表明，AtOR是一种双重靶向蛋白，主要位于黄化子叶的细胞核中。

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