BACKGROUND & AIMS:
:Recent findings suggest a pivotal role for mitochondria-associated hexokinase in the regulation of apoptosis in animal cells. In this study, virus-induced gene silencing (VIGS) of a hexokinase-encoding Hxk1 caused necrotic lesions on leaves, abnormal leaf morphology, and retarded plant growth in Nicotiana benthamiana. Hxk1 was associated with the mitochondria, and this association required the N-terminal membrane anchor. VIGS of Hxk1 reduced the cellular glucose-phosphorylating activity to approximately 31% of control levels without changing the fructose-phosphorylating activity and did not alter hexose phosphate content severely. The affected cells showed programmed cell death (PCD) morphological markers, including nuclear condensation and DNA fragmentation. Similar to animal cell apoptosis, cytochrome c was released into the cytosol and caspase-9- and caspase-3-like proteolytic activities were strongly induced. Furthermore, based on flow cytometry, Arabidopsis thaliana plants overexpressing Arabidopsis HXK1 and HXK2, both of which are predominantly associated with mitochondria, exhibited enhanced resistance to H(2)O(2)- and alpha-picolinic acid-induced PCD. Finally, the addition of recombinant Hxk1 to mitochondria-enriched fractions prevented H(2)O(2)/clotrimazole-induced cytochrome c release and loss of mitochondrial membrane potential. Together, these results show that hexokinase critically regulates the execution of PCD in plant cells, suggesting a link between glucose metabolism and apoptosis.
背景与目标:
:最近的发现表明线粒体相关的己糖激酶在调节动物细胞凋亡中起着关键作用。在这项研究中,编码己糖激酶的Hxk1的病毒诱导的基因沉默(VIGS)导致了本特烟草的叶片坏死性病变,异常的叶片形态并延缓了植物的生长。 Hxk1与线粒体相关联,这种关联需要N端膜锚。 Hxk1的VIGS可以将细胞的葡萄糖磷酸化活性降低至对照水平的约31%,而不会改变果糖磷酸化活性,并且不会严重改变磷酸己糖的含量。受影响的细胞显示出程序性细胞死亡(PCD)形态标志物,包括核浓缩和DNA片段化。与动物细胞凋亡相似,细胞色素c被释放到细胞质中,并强烈诱导了caspase-9-和caspase-3-like蛋白水解活性。此外,基于流式细胞仪,拟南芥植物过表达的拟南芥HXK1和HXK2,这两个主要与线粒体相关联,表现出增强的抗H(2)O(2)-和α-吡啶甲酸诱导的PCD。最后,向线粒体富集的部分中添加重组Hxk1可以防止H(2)O(2)/克霉唑诱导的细胞色素c释放和线粒体膜电位的损失。总之,这些结果表明,己糖激酶严格调节植物细胞中PCD的执行,表明葡萄糖代谢与细胞凋亡之间存在联系。