O'Brien I E, Murray B G, Baguley B C, Morris B A, Ferguson I B
Horticulture and Food Research Institute of New Zealand, Auckland, New Zealand.
Exp Cell Res. 1998 May 25;241(1):46-54. doi: 10.1006/excr.1998.4036.
A large decrease in fluorescence intensity of propidium iodide (PI)-stained nuclei is observed during senescence of plant cells. The phenomenon reflects a decrease in accessibility of DNA to this fluorochrome and is a consequence of chromatin condensation. This decrease is substantially greater than usually found in animal nuclei whose chromatin undergoes condensation, e.g., during differentiation or quiescence. Chromatin condensation was confirmed by analyses of (i) DNA accessibility to DNase I, (ii) histone disassociation induced by HCl, (iii) saturation of binding sites by the PI fluorochrome (iv), and (v) visual inspection by fluorescence and confocal microscopy. The extent of changes revealed by these assays was used to map progressive changes in chromatin condensation which allowed us to identify different stages in an apoptosis-like pathway in plants. The initial step of chromatin condensation which occurred prior to endonucleolytic DNA degradation was detected by fluorescence and confocal microscopy and confirmed by a variety of assays employing flow cytometry. The initial chromatin condensation appears to be a reversible step in the early stage of apoptosis. The loss of reversibility of chromatin condensation observed subsequently may be a critical point in the cascade of apoptotic events, leading to further irreversible changes during apoptosis in plants.
在植物细胞衰老过程中，观察到碘化丙啶（PI）染色细胞核的荧光强度大幅下降。这种现象反映了DNA与这种荧光染料结合能力的降低，是染色质凝聚的结果。这种下降幅度明显大于通常在经历染色质凝聚的动物细胞核中观察到的情况，例如在分化或静止期。通过以下分析证实了染色质凝聚：（i）DNA对DNase I的可及性，（ii）HCl诱导的组蛋白解离，（iii）PI荧光染料对结合位点的饱和情况，（iv）以及通过荧光和共聚焦显微镜进行的目视检查。这些检测所揭示的变化程度被用于描绘染色质凝聚的渐进性变化图谱，这使我们能够识别植物中类似凋亡途径的不同阶段。在核酸内切酶降解DNA之前发生的染色质凝聚的初始步骤，通过荧光和共聚焦显微镜检测到，并通过多种流式细胞术检测方法得到证实。初始的染色质凝聚似乎是凋亡早期的一个可逆步骤。随后观察到的染色质凝聚不可逆性的丧失可能是凋亡事件级联反应中的一个关键点，导致植物凋亡过程中进一步的不可逆变化。
