储存过程中的种子变质是农业和林业生产以及种质保护中的主要问题。我们先前的研究表明,线粒体外膜蛋白电压依赖性阴离子通道(VDAC)参与了榆树(UlmuspumilaL.)种子的受控退化处理(CDT)过程中的程序性细胞死亡(PCD)样活力丧失,但其潜在机制仍不清楚。在这项研究中,我们证明,在榆树种子CDT期间,甘油-3-磷酸脱氢酶(GAPDH)的氧化修饰在VDAC的门调节中起作用。通过生化和细胞学方法和观察转基因材料[拟南芥(拟南芥),Nicotianabenthamiana,和酵母(酿酒酵母)],我们证明了半胱氨酸S-谷胱甘肽化的UpGAPDH1在种子老化过程中与UpVDAC3相互作用,导致线粒体通透性转变和细胞死亡加剧,如线粒体促凋亡因子细胞色素c的泄漏和凋亡核的出现所示。生理测定和电感耦合等离子体质谱(ICP-MS)分析显示GAPDH谷胱甘肽化是由增加的谷胱甘肽介导的,这可能是由于游离金属浓度的增加,特别是Zn。Zn特异性螯合剂TPEN的介绍[(N,N,N\',N'-四(2-吡啶基甲基)乙二胺)]显着延迟种子老化。我们得出的结论是,谷胱甘肽化的UpGAPDH1与UpVDAC3相互作用,并在种子老化过程中充当VDAC门控调节和细胞死亡起始的促凋亡蛋白。
Seed deterioration during storage is a major problem in agricultural and forestry production and for germplasm conservation. Our previous studies have shown that a mitochondrial outer membrane protein VOLTAGE-DEPENDENT ANION CHANNEL (
VDAC) is involved in programmed cell death-like viability loss during the controlled deterioration treatment (CDT) of elm (Ulmus pumila L.) seeds, but its underlying mechanism remains unclear. In this study, we demonstrate that the oxidative modification of GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE (GAPDH) is functioned in the gate regulation of
VDAC during the CDT of elm seeds. Through biochemical and cytological methods and observations of transgenic material [Arabidopsis (Arabidopsis thaliana), Nicotiana benthamiana, and yeast (Saccharomyces cerevisiae)], we demonstrate that cysteine S-glutathionylated UpGAPDH1 interacts with UpVDAC3 during seed aging, which leads to a mitochondrial permeability transition and aggravation of cell death, as indicated by the leakage of the mitochondrial proapoptotic factor cytochrome c and the emergence of apoptotic nucleus. Physiological assays and inductively coupled plasma mass spectrometry analysis revealed that GAPDH glutathionylation is mediated by increased glutathione, which might be caused by increases in the concentrations of free metals, especially Zn. Introduction of the Zn-specific chelator TPEN [(N,N,N\',N\'-Tetrakis (2-pyridylmethyl)ethylenediamine)] significantly delayed seed aging. We conclude that glutathionylated UpGAPDH1 interacts with UpVDAC3 and serves as a proapoptotic protein for
VDAC-gating regulation and cell death initiation during seed aging.