Programmed Cell Death in Plants
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Programmed cell death (PCD) is an important mechanism in development and in defense against pathogens in both animals and plants. PCD may be activated by a variety of environmental or biological cues. Plants have at least two types of programmed cell death. Apoptosis is one form of PCD, some features of which may be evolutionarily conserved in both plants and animals. Cysteine proteases are involved in a key step in animal PCD pathways, and have recently been demonstrated to also be important in plant PCD (Solomon et al., 1999). The authors cultured soybean cells in hydrogen-peroxide to induce PCD and were found to have high levels of specific proteases (e.g. caspases). Intercellular signaling may regulate specific protease inhibitor genes (e.g. cystatin), which in turn, regulate protease activity. Salicylic acid inhibits cystatin expression and may do so under high hydrogen-peroxide concentrations -- effectively promoting PCD. Conversely, jasmonate was found to inhibit PCD. Oxidative stress from hydrogen peroxide may result through mechanical damage of cells by the feeding of herbivores. Solomon et al. (1999) have postulated that downregulation of PCD by jasmonate may occur in cases where there is no survival advantage for PCD (e.g. when there is attack by large herbivores). The morphological changes in tobacco caused by PCD have been described by Mittler et al. (1997). Tobacco plants infected with tobacco mosaic virus (TMV) were used to study the hypersensitive response (HR) activated by PCD. When tobacco plants are grown at 30°C, PCD is inhibited which allows TMV infection to spread. TMV-infected plants were shifted from 30°C to 25°C to induce PCD. Uninfected plants also temperature shifted, and TMV-infected plants grown at 30°C, were used as controls. Also, freeze-thaw induced necrosis was used as a control for cell death not caused by PCD (i.e. cell death not requiring the activation of some protein). Mittler et al. (1997) observed stem and leaf sections using light microscopy, and they found that PCD occurs simultaneously in most leaf cells, but asynchronously in stem cells. Transmission electron microscopy (TEM) was used to visualise the morphology of cells in stem sections of TMV-infected plants and the control plants. In contrast to animal cells, no apoptotic bodies were seen -- the authors suggest this may be due to the cell wall which would prevent engulfment by neighbouring cells. However, vacuolization of the cytoplasm and condensation of chromatin was observed, which are similar to events in apoptosis of animal systems. Also, chloroplasts from TMV-infected cells were found to contain starch granules not present in uninfected cells, necrotic cells, or in cells infected by a bacterial pathogen (Pseudomonas syringae). The degradation of DNA also appears to be a common feature of PCD in plants and animals. In some animal systems, large DNA fragments of 300kb and 50kb are produced during PCD, and 180bp segments indicating internucleosomal fragmentation may also be generated. Mittler et al. (1997) detected 50kb nuclear DNA fragments in both PCD (at 48 hours) and freeze-thaw (in one hour) treatments using field inversion gel electrophoresis (FIGE) and DNA hybridization (with probes specific for nuclear, chloroplast, or mitochondrial DNA). Thus, nuclear DNA degradation may be a response to mechanical injury and can be mediated by PCD in the hypersensitive response. Fragments of 180 kb were not detected, suggesting there is no internucleosomal cleavage in plants during PCD. The chloroplast genome normally exists in multimeric forms, but levels of the monomeric form were found by Mittler et al. (1997) to increase early on during TMV-induced PCD. One way to test whether plants and animals share common PCD pathways would be to isolate nuclei from animal cells, transfer them into the cytosol of plant cells and induce PCD. Zhao et al (1999) developed a cell free system using mouse liver nuclei and carrot cytosol. Cytochrome c is required to activate caspases, which generate a PCD response. Over a period of four hours upon addition of cytochrome c to their system, Zhao et al. (1999) detected chromatin condensation and the subsequent formation of apoptotic bodies that were extruded into the cytosol. Zhao et al. detected 3'-OH DNA strand breaks yielding a DNA ladder, using the TUNEL (transferase-mediated dUTP-digoxigenin nick end labeling) assay. They also investigated the presence and influence of cysteine proteases using caspase inhibitors. Zhao et al (1999) have indicated that research into the mechanisms of PCD and apoptosis in plants is far behind that in animals. In general, it appears that plants and animals do share some common pathways in programmed cell death. However, some features may be unique to plants.
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