利用快速共聚焦显微镜观察花粉管中线粒体的分布及其动态变化

线粒体是真核细胞中高度动态变化的一种细胞器。但目前有关植物细胞,尤其是花粉管中线粒体的分布及其动态变化的信息还比较少。本文应用Zeiss 5 live快速共聚焦显微镜结合线粒体荧光探针Mitotracker Green对百合花粉管中线粒体的分布及其动态变化进行了观察和测定。结果显示,正常培养的百合花粉管中线粒体呈倒喷泉式移动,即花粉管基部的线粒体移动到亚顶端后即发生回流,因此花粉管顶端锥形区域内很少观察到线粒体的存在。对单个线粒体进行跟踪分析结果表明花粉管中的线粒体分为快速运动和锚定状态两种。快速移动的线粒体在花粉管两侧质膜下及花粉管中央沿着与花粉管长轴平行的方向运动,在花粉管亚顶端则沿着一定曲线移动;锚定在细胞质中的线粒体则随着胞质环流而被动移动。低浓度的微丝骨架抑制剂Jas处理时间依赖性地引起花粉管亚顶端的线粒体逐渐前移,并最终充满整个花粉管顶端,同时花粉管中快速移动线粒体的比例逐渐减少。上述结果表明,花粉管中的线粒体主要沿着微丝骨架进行快速移动,而花粉管亚顶端精细微丝组成的领区(Collar)在花粉管线粒体的分布和动态变化中起重要作用。

Visualization the distribution and dynamics of mitochondria in pollen tubes by real-time laser scanning confocal microscope

Mitochondria are vital organelles involved in various variety of fundamental functions ranging from the biosynthesis of ATP through the programmed cell death. However, the present data are apparently insufficient to provide complete knowledge of the distribution and dynamics of mitochondria, especially in plant cell. In the present study, mitochondria in Lilium longiflorum was stained with Mitotracker Green and visualized with Zeiss 5 live real-time laser scanning confocal microscope. It was revealed that mitochondria in the control tubes streams are in a reverse fountain pattern. In detail, in Lilium longiflorum pollen tubes, mitochondria move beneath the plasma membrane toward the subapex where they turn and move away from center of the tube toward the base, mitochondria were therefore rarely observed at the apex of pollen tube. Monitoring individual mitochondria indicated that their motion can be mainly classified into two categories: some move rapidly with preferentially axial orientation both in the center and beneath the plasma membrane of pollen tube, but gradually change their direction following a semicircular track at the subapex. The others were in Brownian motion, passively change their position promoted by cytoplasmic streaming. Application of Jas, which disrupts microfilaments, resulted in the extension of mitochondria into the extreme apex in a time dependent manner. Gradually, the amount of rapid moving mitochondria reduced. All of these data enable us to conclude that rapid movement of mitochondria mainly depend on microfilaments, and the "collar", a distinct arrangement of mostly net-axially orientated AFs at the subapex, may has vital function in mitochondria distribution in pollen tubes.