Table_2_AhABI4s Negatively Regulate Salt-Stress Response in Peanut.DOCX

Soil salinity is one of the major factors that limit the area of cultivable land and yield potential of crops. The ability of salt tolerance varies with plant species. Peanut (Arachis hypogaea L.) is a moderately salt-sensitive and economically important crop, however, their biological processes involved in salt-stress response remain unclear. In this study, we investigated the role of A. hypogaea L. ABSCISIC ACID INSENSITIVE 4s (AhABI4s) in salt tolerance and elucidated its mode of action in peanuts. The results showed that the downregulation of AhABI4s via whole plant virus-induced gene silencing has enhanced the survival rate, biomass accumulation, and root/shoot ratio of peanut seedlings in response to salt-stress. Transcriptomics, quantitative proteomics, and phosphoproteomic analyses were performed using AhABI4s-silenced and Mock plants. The expression pattern of 15,247 genes, 1,900 proteins, and 2,620 phosphorylation sites were affected by silencing of AhABI4s in peanut leaf and root after sodium chloride (NaCl) treatment. Among them, 63 potential downstream target genes of ABI4 changed consistently at both transcription and translation levels, and the protein/phosphorylation levels of 31 ion transporters/channels were also affected. Electrophoretic mobility shift assays (EMSA) showed that ABI4 was able to bind to the promoters of HSP70, fructokinase (FRK), and pyruvate kinase (PK) coding genes in vitro. In addition, we also detected a binding preference of AhABI4 for CACT(G/T)GCA motif in the promoters of down-regulated genes in peanut leaf. Collectively, the potential downstream targets which were regulated at the levels of transcription and translation, binding preference, and in vivo phosphorylation sites that had been revealed in this study will provide new insight into the AhABI4s-mediated salt tolerance regulation mechanism in peanuts.

土壤盐度是限制可耕地面积与作物产量潜力的核心影响因素之一。不同植物物种的耐盐能力存在显著差异。花生（Arachis hypogaea L.）属于中度盐敏感型经济作物，但其参与盐胁迫应答的生物学过程仍有待阐明。本研究探究了花生脱落酸不敏感4基因（AhABI4s，全称ABSCISIC ACID INSENSITIVE 4）在耐盐性中的功能，并阐明了其在花生体内的作用模式。研究结果显示，通过整株病毒诱导基因沉默下调AhABI4s的表达，可显著提升盐胁迫下花生幼苗的存活率、生物量积累量以及根冠比。研究团队对AhABI4s沉默植株与空白对照（Mock）植株开展了转录组学、定量蛋白质组学以及磷酸化蛋白质组学分析。经氯化钠（NaCl）处理后，花生叶片与根系中共有15247个基因、1900个蛋白质以及2620个磷酸化位点的表达模式因AhABI4s沉默发生改变。其中，63个ABI4潜在下游靶基因在转录与翻译水平上的变化趋势一致；另有31个离子转运蛋白/通道的蛋白质水平及磷酸化水平受到调控影响。电泳迁移率变动分析（Electrophoretic Mobility Shift Assay, EMSA）结果表明，ABI4可在体外结合热休克蛋白70（HSP70）、果糖激酶（fructokinase, FRK）以及丙酮酸激酶（pyruvate kinase, PK）的编码基因启动子区域。此外，本研究还发现，AhABI4对花生叶片中下调基因启动子区域的CACT(G/T)GCA基序具有结合偏好性。综上，本研究揭示的转录与翻译水平调控的潜在下游靶基因、结合偏好性以及体内磷酸化位点，将为解析花生中AhABI4s介导的耐盐调控机制提供全新的研究视角。