College of Horticulture, Northwest A&F University, Yangling, 712100, China.
Department of Horticulture, The University of Haripur, Haripur, 22620, Pakistan.
Plant J. 2024 Mar;117(5):1377-1391. doi: 10.1111/tpj.16568. Epub 2023 Nov 28.
Plant NAC transcription factors play a crucial role in enhancing cold stress tolerance, yet the precise molecular mechanisms underlying cold stress remain elusive. In this study, we identified and characterized CaNAC035, an NAC transcription factor isolated from pepper (Capsicum annuum) leaves. We observed that the expression of the CaNAC035 gene is induced by both cold and abscisic acid (ABA) treatments, and we elucidated its positive regulatory role in cold stress tolerance. Overexpression of CaNAC035 resulted in enhanced cold stress tolerance, while knockdown of CaNAC035 significantly reduced resistance to cold stress. Additionally, we discovered that CaSnRK2.4, a SnRK2 protein, plays an essential role in cold tolerance. In this study, we demonstrated that CaSnRK2.4 physically interacts with and phosphorylates CaNAC035 both in vitro and in vivo. Moreover, the expression of two ABA biosynthesis-related genes, CaAAO3 and CaNCED3, was significantly upregulated in the CaNAC035-overexpressing transgenic pepper lines. Yeast one-hybrid, Dual Luciferase, and electrophoretic mobility shift assays provided evidence that CaNAC035 binds to the promoter regions of both CaAAO3 and CaNCED3 in vivo and in vitro. Notably, treatment of transgenic pepper with 50 μm Fluridone (Flu) enhanced cold tolerance, while the exogenous application of ABA at a concentration of 10 μm noticeably reduced cold tolerance in the virus-induced gene silencing line. Overall, our findings highlight the involvement of CaNAC035 in the cold response of pepper and provide valuable insights into the molecular mechanisms underlying cold tolerance. These results offer promising prospects for molecular breeding strategies aimed at improving cold tolerance in pepper and other crops.
植物 NAC 转录因子在增强植物的抗冷性方面发挥着至关重要的作用，但冷胁迫的精确分子机制仍难以捉摸。在本研究中，我们从辣椒叶片中分离并鉴定了一个 NAC 转录因子 CaNAC035。我们观察到 CaNAC035 基因的表达受冷胁迫和脱落酸（ABA）处理的诱导，并阐明了其在冷胁迫耐受性中的正调控作用。过表达 CaNAC035 可增强冷胁迫耐受性，而 CaNAC035 的敲低则显著降低了对冷胁迫的抗性。此外，我们发现 CaSnRK2.4，一种 SnRK2 蛋白，在耐冷性中起着重要作用。在本研究中，我们证明 CaSnRK2.4 在体内和体外均与 CaNAC035 相互作用并磷酸化 CaNAC035。此外，两个 ABA 生物合成相关基因 CaAAO3 和 CaNCED3 的表达在 CaNAC035 过表达的转基因辣椒株系中显著上调。酵母单杂交、双荧光素酶报告基因和电泳迁移率变动分析实验提供了证据表明 CaNAC035 在体内和体外均可与 CaAAO3 和 CaNCED3 的启动子区域结合。值得注意的是，用 50 μM Fluridone（Flu）处理转基因辣椒可增强其耐冷性，而用 10 μM ABA 处理病毒诱导基因沉默系则明显降低了其耐冷性。总之，我们的研究结果表明 CaNAC035 参与了辣椒的冷响应，并为冷胁迫耐受性的分子机制提供了有价值的见解。这些结果为旨在提高辣椒和其他作物耐冷性的分子育种策略提供了有前景的思路。
