Major changes in chromatin condensation suggest the presence of an apoptotic pathway in plant cells.

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.

[1]  R. Gardner,et al.  Flow cytometric determination of genome size in Pinus , 1996 .

[2]  Z. Darżynkiewicz,et al.  Changes in nuclear chromatin related to apoptosis or necrosis induced by the DNA topoisomerase II inhibitor fostriecin in MOLT-4 and HL-60 cells are revealed by altered DNA sensitivity to denaturation. , 1992, Experimental cell research.

[3]  E. Macrae,et al.  Xyloglucan endotransglycosylase activity during fruit development and ripening of apple and kiwifruit , 1996 .

[4]  D. Galbraith,et al.  Rapid Flow Cytometric Analysis of the Cell Cycle in Intact Plant Tissues , 1983, Science.

[5]  Z. Darżynkiewicz,et al.  Flow cytometric detection of apoptosis: comparison of the assays of in situ DNA degradation and chromatin changes. , 1994, Cytometry.

[6]  Z. Darżynkiewicz,et al.  Features of apoptotic cells measured by flow cytometry. , 1992, Cytometry.

[7]  D. Ryerson,et al.  Cleavage of Nuclear DNA into Oligonucleosomal Fragments during Cell Death Induced by Fungal Infection or by Abiotic Treatments. , 1996, The Plant cell.

[8]  Alan M. Jones,et al.  Logjam at the Styx: Programmed cell death in plants , 1996 .

[9]  T. Lindmo,et al.  Flow cytometry and sorting , 1979 .

[10]  H. Crissman,et al.  Use of flow microfluorometry in detailed analysis of effects of chemical agents on cell cycle progression. , 1972, Cancer research.

[11]  D L Evans,et al.  Analysis and discrimination of necrosis and apoptosis (programmed cell death) by multiparameter flow cytometry. , 1992, Biochimica et biophysica acta.

[12]  Z. Darżynkiewicz,et al.  Detection of DNA strand breaks in individual apoptotic cells by the in situ terminal deoxynucleotidyl transferase and nick translation assays. , 1993, Cancer research.

[13]  R. Bostock,et al.  Apoptosis: A Functional Paradigm for Programmed Plant Cell Death Induced by a Host-Selective Phytotoxin and Invoked during Development. , 1996, The Plant cell.

[14]  C. Reutelingsperger,et al.  Annexin-V and TUNEL use in monitoring the progression of apoptosis in plants. , 1997, Cytometry.

[15]  R. Newton,et al.  Genome size and environmental factors in the genus Pinus , 1993 .

[16]  A. Rayburn,et al.  Flow cytometric probing of chromatin condensation in maize diploid nuclei , 1994 .

[17]  Z. Darżynkiewicz,et al.  Altered Sensitivity of DNA in Situ to Denaturation in Apoptotic Cells a , 1993, Annals of the New York Academy of Sciences.

[18]  J. Miksche,et al.  DNA CONTENT AND HETEROCHROMATIN VARIATIONS IN VARIOUS TISSUES OF PEANUT (ARACHIS HYPOGAEA) , 1982 .