Tobacco Transcription Factor NtbHLH123 Confers Tolerance to Cold Stress by Regulating the NtCBF Pathway and Reactive Oxygen Species Homeostasis

Abstract

Cold stress is a major environmental factor that impairs plant growth and development, geographic distribution, and crop productivity. The C-repeat binding factor (CBF) regulatory pathway has an essential role in response to cold stress. Here, we characterized a bHLH transcription factor from <i>Nicotiana tabacum</i>, <i>NtbHLH123</i>, in response to cold stress (4°C). Overexpression of <i>NtbHLH123</i> enhanced cold tolerance in transgenic tobacco plants. Based on yeast one-hybrid, chromatin immunoprecipitation PCR, and transient expression analysis assays, <i>NtbHLH123</i> binds directly to the G-box/E-box motifs in the promoter of the <i>NtCBF</i> genes and positively regulates their expression. Furthermore, <i>NtbHLH123</i>-overexpressing plants showed lower electrolyte leakage, reduced malondialdehyde contents, H₂O₂ and reactive oxygen species (ROS) accumulation under cold stress, which contributed to alleviating oxidative damage to the cell membrane after cold stress treatment. And <i>NtbHLH123</i> increased stress tolerance by improving the expression of a number of abiotic stress-responsive genes to mediate the ROS scavenging ability and other stress tolerance pathways. Taken together, we present a model suggesting that <i>NtbHLH123</i> is a transcriptional activator that functions as a positive regulator of cold tolerance by activating <i>NtCBF</i>, ROS scavenging-related, and stress-responsive genes.