大叶桉挥发物对蚕豆细胞毒性效应及信号调节

Toxic effects and signal regulation of Eucalyptus robusta volatiles on Vicia faba cells

关键词:  大叶桉, 挥发物, 化感作用, 毒性效应, 信号调节

Key words:  Eucalyptus robusta, volatiles, allelopathy, toxic effects, signal regulation

摘要:

为探讨大叶桉(Eucalyptus robusta)化感作用的细胞学机制,该研究以大叶桉挥发油及其主要成分α-蒎烯和桉油精为供体,以蚕豆(Vicia faba )的根细胞和叶保卫细胞为靶标,运用显微技术、细胞化学技术和qRT-PCR技术,研究了大叶桉挥发物的毒性效应。结果表明:(1)在大叶桉挥发物作用下,蚕豆幼根生长受抑制并表现为时间-浓度依赖效应,其化感效应强弱由大到小依次为挥发油、α-蒎烯和桉油精。(2)蚕豆根边缘细胞活性降低,分生区细胞微核率升高,有丝分裂指数下降且大部分细胞的细胞周期被阻滞在分裂前期。(3)蚕豆叶保卫细胞内NADPH氧化酶活性升高,活性氧(reactive oxygen species, ROS)爆发,微丝聚合,气孔开度下降; 叶保卫细胞的核畸变率升高,细胞活性降低甚至发生caspase依赖性细胞凋亡,而Ca²⁺通道阻断剂(LaCl₃)、活性氧清除剂(AsA)和硝酸还原酶抑制剂(NaN₃)均可显著提高保卫细胞存活率,说明大叶桉挥发物改变了信号分子Ca²⁺、ROS和NO的信号调节。综上表明,大叶桉挥发物的细胞毒性和遗传毒性改变了受体细胞的信号转导途径,诱发了细胞遗传畸变,导致受体根边缘细胞保护功能障碍和气孔运动障碍,从而影响受体根系生长和光合作用,最终导致受体生长受阻。该研究结果为大叶桉种植区的科学种植和管理提供了理论依据。

Abstract:

To understand the cytological mechanism of allelopathy of Eucalyptus robusta, the toxic effects of volatiles from E. robusta were studied by microscopic, cytochemical and qRT-PCR techniques, taking volatile oil from E. robusta and its main components α-pinene and eucalyptol as donors, and using root cells and leaf guard cells of Vicia faba as targets. The results were as follows:(1)The growth of radicle of V. faba were exhibited and showed a time-concentration dependent effects under the treatments of the volatiles of Eucalyptus robusta. The allelopathic effects were volatile oil, α-pinene and eucalyptol in descending order.(2)When Vicia faba roots were exposed to Eucalyptus robusta volatiles, the activity of root border cells decreased, the micronucleus rate of cells in the meristematic zone increased, mitotic index decreased, and the cell cycle of most cells was arrested in the prophase of division.(3)Under the action of the volatiles of E. robusta, the activity of NADPH oxidase increased, reactive oxygen species(ROS)burst in leaf guard cells of Vicia faba, microfilament polymerization, and stomatal aperture decreased. At the same time, the leaf epidermis strip of V. faba was treated with Eucalyptus robusta volatiles, the nuclear distortion rate of leaf guard cells increased. Moreover, the treatment of E. robusta volatiles led to the decrease of guard cell activity and caspase-dependent apoptosis in Vicia faba. However, the guard cell survival rates increased when the leaf epidermis strips of V. faba were exposed to volatiles from Eucalyptus robusta combined with different concentrations of Ca²⁺channel blocker(LaCl₃), ROS scavenger ascorbic acid(AsA), and nitrate reductase inhibitors(NaN₃), which indicated that the volatiles of E. robusta changed the signal regulation of Ca²⁺, ROS and NO. These results suggested that the cytotoxicity and genotoxicity of the volatiles of E. robusta altered the signal transduction pathway of the receptor cells, induced the genetic aberration of the root tip cells, then led to the dysfunction of protective function and stomatal movement of the receptor root border cells, which affected the root growth and photosynthesis of the receptor, and ultimately hindered the growth of receptor. The results provide a theoretical basis for scientific planting and management of E. robusta planting area.