Table_1_MdbHLH130, an Apple bHLH Transcription Factor, Confers Water Stress Resistance by Regulating Stomatal Closure and ROS Homeostasis in Transgenic Tobacco.docx

Drought is a major environmental factor that significantly limits crop yield and quality worldwide. Basic helix-loop-helix (bHLH) transcription factors have been reported to participate in the regulation of various abiotic stresses. In this study, a bHLH transcription factor in apple, MdbHLH130, which contains a highly conserved bHLH domain, was isolated and characterized. qRT-PCR and PMdbHLH130::GUS analyses showed that MdbHLH130 was notably induced in response to dehydration stress. Compared with the wild-type (WT), transgenic apple calli overexpressing MdbHLH130 displayed greater resistance to PEG6000 treatment. In contrast, the MdbHLH130-Anti lines were more sensitive to PEG6000 treatment than WT. Moreover, ectopic expression of MdbHLH130 in tobacco improved tolerance to water deficit stress, and plants exhibited higher germination rates and survival rates, longer roots, and lower ABA-induced stomatal closure and leaf water loss than the WT control. Furthermore, overexpression of MdbHLH130 in tobacco also led to lower electrolyte leakage, malondialdehyde contents, and reactive oxygen species (ROS) accumulation and upregulation of the expression of some ROS-scavenging and stress-responsive genes under water deficit stress. In addition, MdbHLH130 transgenic tobacco plants exhibited improved tolerance to oxidative stress compared with WT. In conclusion, these results indicate that MdbHLH130 acts as a positive regulator of water stress responses through modulating stomatal closure and ROS-scavenging in tobacco.

干旱是全球范围内显著限制作物产量与品质的主要环境胁迫因子。基本螺旋-环-螺旋（basic helix-loop-helix, bHLH）转录因子已有研究表明可参与调控多种非生物胁迫响应。本研究从苹果中分离得到一个携带高度保守bHLH结构域的bHLH转录因子MdbHLH130，并对其进行了功能鉴定与表征。qRT-PCR与PMdbHLH130::GUS分析结果显示，MdbHLH130的表达显著受脱水胁迫诱导。与野生型（wild-type, WT）相比，过表达MdbHLH130的转基因苹果愈伤组织对PEG6000处理表现出更强的抗性；反之，MdbHLH130反义表达株系相较于野生型，对PEG6000处理更为敏感。此外，在烟草中异位表达MdbHLH130可提升植株对水分亏缺胁迫的耐受性：与野生型对照相比，转基因植株的发芽率与存活率更高、根系更长，且ABA诱导的气孔关闭程度与叶片水分流失率更低。进一步研究发现，水分亏缺胁迫下，烟草中过表达MdbHLH130还可降低电解质渗漏率、丙二醛含量与活性氧（reactive oxygen species, ROS）积累量，并上调部分ROS清除基因与胁迫响应基因的表达水平。除此之外，与野生型相比，MdbHLH130转基因烟草对氧化胁迫的耐受性亦显著提升。综上，上述结果表明，MdbHLH130可通过调控烟草的气孔关闭与ROS清除通路，作为正调控因子参与水分胁迫响应。