NaCl胁迫下平邑甜茶根系线粒体特性和细胞死亡特征

Characteristics of mitochondria and cell death in roots of Malus hupehensis var. pingyiensis under NaCl stress

以平邑甜茶(Malushupehensisvar.pingyiensis)实生幼苗为试验材料,研究NaCl浇灌后根系线粒体H2O2含量、膜电位(△ψm)和根系ATP含量的变化以及细胞死亡特征。结果表明,根系线粒体H2O2含量在0.085mol·L–1NaCl处理的第1–6天逐渐降低,在第6–15天则快速上升;线粒体△ψm在0.085mol·L–1NaCl处理的15天内一直呈下降趋势,在第6–15天下降速度明显加快;根系ATP含量在0.085mol·L–1NaCl处理的15天内始终低于对照,但保持在一个较稳定的范围内。TUNEL原位末端标记试验显示,0.085mol·L–1NaCl处理的第9天,根系石蜡组织切片上的阳性反应斑点明显增多,到第15天时阳性反应斑点密集成片,表明细胞核DNA发生了细胞程序性死亡的特征性断裂。根系中细胞程序性死亡关键酶类caspase3/7活性在0.085mol·L–1NaCl处理的第1–6天处于较低水平,其活性在第6–15天成倍上升。这些结果表明,0.085mol·L–1NaCl处理6–15天能诱导平邑甜茶根细胞发生程序性死亡,而且线粒体特性的变化与根系细胞程序性死亡密切相关。

Aims Malus hupehensis var. pingyiensis with strong ability for apomixes is unique to China and is tolerant of waterlogged conditions and salt. There is little difference among its seedlings because of apomixes. Malus hupe-hensis var. pingyiensis is widely used as stocks of apple cultivars and ornamental apples and has important applied value in apple production and research. Our objective was to determine the response characteristic of cells and mitochondria in M. hupehensis var. pingyiensis roots under high saline conditions. Methods Pot seedlings with at least seven mature leaves were used in this experiment. In order to avoid irre-versible damage to roots caused by sudden exposure to high level saline condition, we first used a weak saline solution （0.051 mol·L–1 NaCl） to water seedlings every two days, for a total of three times. Then strong saline so-lution （0.085 mol·L–1 NaCl） was used to water seedlings. Tap water was used for control seedlings. H2O2 content, membrane potential of mitochondria （Δψm）, ATP content, terminal deoxynucleotidyl transferase mediated dUTP nick end labelling （TUNEL） and caspase3/7 activity were determined every two days from the third day of 0.085 mol·L–1 NaCl treatment, altogether five times. Important findings Δψm of roots declined slowly in the first six days of 0.085 mol·L–1 NaCl treatment and thereafter declined rapidly. The H2O2 content in mitochondria decreased slowly in the first six days and then ac-cumulated quickly. ATP content in roots remained constant during the treatment of 0.085 mol·L–1 NaCl, but was low compared with that in control seedlings. The results of TUNEL assay showed that root paraffin slices dis-played distinct positive-reaction spots that became more numerous in the root paraffin slice of the ninth day. Cas-pase3/7 activity was examined with a detection kit; it remained at a low level for the first six days of 0.085 mol·L–1 NaCl treatment, and thereafter increased sharply. Results indicated that programmed cell death could be induced in roots of M. hupehensis var. pingyiensis by 0.085 mol·L–1 NaCl treatment, and this was closely related to the change of mitochondrial characteristics.