The Photorespiratory BOU Gene Mutation Alters Sulfur Assimilation and Its Crosstalk With Carbon and Nitrogen Metabolism in Arabidopsis thaliana. Arabidopsis thaliana

This study was aimed at elucidating the significance of photorespiratory serine (Ser) production for cysteine (Cys) biosynthesis. For this purpose, sulfur (S) metabolism and its crosstalk with nitrogen (N) and carbon (C) metabolism were analyzed in wildtype Arabidopsis and its photorespiratory bou-2 mutant with impaired glycine decarboxylase (GDC) activity. Foliar glycine and Ser contents were enhanced in the mutant at day and night. The high Ser levels in the mutant cannot be explained by transcript abundances of genes of the photorespiratory pathway or two alternative pathways of Ser biosynthesis. Despite enhanced foliar Ser, reduced GDC activity mediated a decline in sulfur flux into major sulfur pools in the mutant, as a result of deregulation of genes of sulfur reduction and assimilation. Still, foliar Cys and glutathione contents in the mutant were enhanced. The use of Cys for methionine and glucosinolates synthesis was reduced in the mutant. Reduced GDC activity in the mutant downregulated Calvin Cycle and nitrogen assimilation genes, upregulated key enzymes of glycolysis and the tricarboxylic acid (TCA) pathway and modified accumulation of sugars and TCA intermediates. Thus, photorespiratory Ser production can be replaced by other metabolic Ser sources, but this replacement deregulates the cross-talk between S, N, and C metabolism. Overall design: Arabidopsis thaliana plants [Wildtype Col-0 and a mutant line in Col-0 background, bou-2 (At5g46800) were grown in biological triplicates under high CO2 (3000 ppm) for 35 days, 12 hr light / 12 hr dark and then shifted to ambient CO2 24 hours before harvesting. Samples for RNASeq were taken two hours before ("end of night", EON) and two hours after ("beginning of day", BOD) the onset of light.

本研究旨在阐明光呼吸丝氨酸（Ser）合成对半胱氨酸（Cys）生物合成的重要意义。为此，我们以野生型拟南芥及其甘氨酸脱羧酶（GDC）活性受损的光呼吸突变体bou-2为研究材料，分析了硫（S）代谢及其与氮（N）、碳（C）代谢的交叉串扰关系。该突变体的叶片甘氨酸与丝氨酸含量在昼夜周期中均显著升高。突变体内高水平的丝氨酸无法通过光呼吸通路或两条替代性丝氨酸合成通路的基因转录本丰度加以解释。尽管叶片丝氨酸含量升高，但由于硫还原与同化相关基因的调控失调，GDC活性降低导致突变体中硫流向主要硫库的通量出现下降。即便如此，突变体的叶片半胱氨酸与谷胱甘肽含量仍有所提升。突变体中用于合成甲硫氨酸与硫代葡萄糖苷的半胱氨酸用量有所降低。突变体中降低的GDC活性会下调卡尔文循环与氮同化相关基因的表达，上调糖酵解与三羧酸（TCA）循环的关键酶，并改变糖类与TCA循环中间产物的积累模式。综上，光呼吸丝氨酸的合成可由其他代谢性丝氨酸来源替代，但该替代过程会破坏硫、氮与碳代谢之间的交叉串扰调控网络。本研究的整体实验设计如下：将拟南芥（Arabidopsis thaliana）哥伦比亚野生型（Col-0）以及Col-0遗传背景下的bou-2突变体（At5g46800）进行三次生物学重复培养，先在高二氧化碳（3000 ppm）环境下以12小时光照/12小时黑暗的周期培养35天，于收获前24小时转移至正常二氧化碳环境后收获样本。RNA测序（RNASeq）的采集时点分别为光照开始前两小时（夜间结束时，EON）与光照开始后两小时（日间起始时，BOD）。