Table_4_Proteomics reveals the significance of vacuole Pi transporter in the adaptability of Brassica napus to Pi deprivation.xlsx

Vacuolar Pi transporters (VPTs) have recently been identified as important regulators of cellular Pi status in Arabidopsis thaliana and Oryza sativa. In the oil crop Brassica napus, BnA09PHT5;1a and BnC09PHT5;1a are two homologs of AtPHT5;1, the vacuolar Pi influx transporter in Arabidopsis. Here, we show that Pi deficiency induces the transcription of both homologs of PHT5;1a genes in B. napus leaves. Brassica PHT5;1a double mutants (DM) had smaller shoots and higher cellular Pi concentrations than wild-type (WT, Westar 10), suggesting the potential role of BnPHT5;1a in modulating cellular Pi status in B. napus. A proteomic analysis was performed to estimate the role of BnPHT5;1a in Pi fluctuation. Results show that Pi deprivation disturbs the abundance of proteins in the physiological processes involved in carbohydrate metabolism, response to stimulus and stress in B. napus, while disruption of BnPHT5;1a genes may exacerbate these processes. Besides, the processes of cell redox homeostasis, lipid metabolic and proton transmembrane transport are supposed to be unbalanced in BnPHT5;1a DM under the -Pi condition. Noteworthy, disruption of BnPHT5;1a genes severely alters the abundance of proteins related to ATP biosynthesis, and proton/inorganic cation transmembrane under normal Pi condition, which might contribute to B. napus growth limitations. Additionally, seven new protein markers of Pi homeostasis are identified in B. napus. Taken together, this study characterizes the important regulatory role of BnPHT5;1a genes as vacuolar Pi influx transporters in Pi homeostasis in B. napus.

液泡无机磷酸盐转运蛋白（Vacuolar Pi Transporters, VPTs）近期被证实为拟南芥（Arabidopsis thaliana）与水稻（Oryza sativa）细胞无机磷酸盐（Pi）稳态的重要调控因子。在油料作物甘蓝型油菜（Brassica napus）中，BnA09PHT5;1a与BnC09PHT5;1a是拟南芥液泡Pi内流转运蛋白AtPHT5;1的两个同源基因。本研究发现，Pi匮乏会诱导甘蓝型油菜叶片中PHT5;1a两个同源基因的转录。甘蓝型油菜BnPHT5;1a双突变体（DM）的地上部较野生型（WT，Westar 10）更小，且细胞内Pi浓度更高，表明BnPHT5;1a可能参与调控甘蓝型油菜的细胞Pi稳态。为探究BnPHT5;1a在Pi波动中的作用，本研究开展了蛋白质组学分析。结果显示，Pi剥夺会干扰甘蓝型油菜中参与碳水化合物代谢、刺激响应与逆境响应生理过程的蛋白丰度，而BnPHT5;1a基因的敲除可能会加剧这些变化。此外，在-Pi（Pi匮乏）条件下，甘蓝型油菜BnPHT5;1a双突变体的细胞氧化还原稳态、脂质代谢以及质子跨膜运输过程可能出现失衡。值得注意的是，在正常Pi条件下，BnPHT5;1a基因的敲除会显著改变与ATP生物合成、质子/无机阳离子跨膜运输相关的蛋白丰度，这可能是导致甘蓝型油菜生长受限的原因。此外，本研究还在甘蓝型油菜中鉴定出7个新的Pi稳态蛋白标志物。综上，本研究阐明了BnPHT5;1a基因作为液泡Pi内流转运蛋白，在甘蓝型油菜Pi稳态调控中的重要作用。