The WHIRLY (WHY) DNA/RNA binding proteins fulfil multiple but poorly characterised functions in leaf development. Here, we show that WHY1 transcript levels were highest in the bases of 7-day old barley leaves. Immunogold labelling revealed that the WHY1 protein was more abundant in the nuclei than the proplastids of the leaf bases. To identify transcripts associated with leaf development we conducted hierarchical clustering of differentially abundant transcripts along the developmental gradient of wild-type leaves. Similarly, metabolite profiling was employed to identify metabolites exhibiting a developmental gradient. A comparative analysis of transcripts and metabolites in barley lines (W1–1 and W1–7) lacking WHY1, which show delayed greening compared with the wild type revealed that the transcript profile of leaf development was largely unchanged in W1–1 and W1–7 leaves. However, there were differences in levels of several transcripts encoding transcription factors associated with chloroplast development. These include a barley homologue of the Arabidopsis GATA transcription factor that regulates stomatal development, greening and chloroplast development, NAC1; two transcripts with similarity to Arabidopsis GLK1 and two transcripts encoding ARF transcriptions factors with functions in leaf morphogenesis and development. Chloroplast proteins were less abundant in the W1–1 and W1–7 leaves than the wild type. The levels of tricarboxylic acid cycle metabolites and GABA were significantly lower in WHY1 knockdown leaves than the wild type. This study provides evidence that WHY1 is localised in the nuclei of leaf bases, contributing the regulation of nuclear-encoded transcripts that regulate chloroplast development.

中文翻译：

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WHIRLY1 在细胞核中发挥调节大麦叶发育和相关代谢物谱的功能

WHIRLY (WHY) DNA/RNA 结合蛋白在叶片发育中实现了多种但特征不佳的功能。在这里，我们显示 WHY1 转录水平在 7 天龄大麦叶的基部中最高。免疫金标记显示，WHY1 蛋白在细胞核中比叶基部的前质体更丰富。为了识别与叶片发育相关的转录本，我们对沿野生型叶片发育梯度的差异丰富的转录本进行了层次聚类。类似地，代谢物分析用于鉴定表现出发育梯度的代谢物。缺乏 WHY1 的大麦系（W1-1 和 W1-7）中转录物和代谢物的比较分析，与野生型相比，显示延迟绿化表明叶片发育的转录谱在 W1-1 和 W1-7 叶片中基本没有变化。然而，编码与叶绿体发育相关的转录因子的几种转录物的水平存在差异。其中包括调节气孔发育、绿化和叶绿体发育的拟南芥 GATA 转录因子的大麦同系物 NAC1；两个与拟南芥 GLK1 相似的转录本和两个编码在叶片形态发生和发育中起作用的 ARF 转录因子的转录本。W1-1 和 W1-7 叶片中的叶绿体蛋白含量低于野生型。WHY1敲低叶片中三羧酸循环代谢物和GABA的水平显着低于野生型。