Plant programmed cell death, ethylene and flower senescence

Programmed cell death (PCD) applies to cell death that is part of the normal life of multicellular organisms. PCD is found throughout the animal and plant kingdoms; it is an active process in which a cell suicide pathway is activated resulting in controlled disassembly of the cell. Most cases of PCD described in animal systems take the form of apoptosis, a cell death process characterised by specific features such as cell shrinkage, blebbing of the plasma membrane, condensation and fragmentation of the nucleus and internucleosomal cleavage of DNA. The final stage of apoptosis is the fragmentation of the cell into cellular debris-containing vesicles called `apoptotic bodies` that are being phagocytosed by other cells. A specific class of cell death- associated cystein proteases (caspases) has been identified. Generally, apoptotic cell death involves a sequence of caspase activation events in which initiator caspases activate down-stream executioner caspases that process a variety of target proteins eventually leading to the apoptotic phenotype. The occurrence of hallmarks of animal apoptosis was studied in tomato cells treated with the anticancer drug and inducer of apoptosis, camptothecin (cpt). It was shown that cpt-induced cell death is accompanied by nuclear condensation, the appearance of TUNEL-positive nuclei, DNA laddering and formation of DNA-containing (apoptotic) bodies and was greatly inhibited by inhibitors of animal caspases. Together the results indicate that cpt induced a cell death pathway with similarities to caspase-mediated (apoptotic) cell death in animal systems. We used cpt-treated cells to study the possible involvement of ethylene in cell death. Treatment of the cells with relatively high concentrations of ethylene did not have any effect on viability of the cells. However, when ethylene was applied in combination with cpt, a significant increase in cell death was observed as compared to cpt treatment alone. Experiments with inhibitors of ethylene production or ethylene action showed that ethylene is an essential factor mediating cpt-induced cell death. Flower senescence is accompanied by rapid death of large numbers of cells. In situ DNA degradation was studied in gypsophila petals using TUNEL. We showed that TUNEL positive nuclei appear well before the onset of the increase in ethylene production and visible signs of senescence. The role of PCD in flower senescence is discussed

Comparison of micro-array profiling in senescing iris and carnation flowers

Gene expression profiles of cut Iris and carnation flowers were studied using cDNA microarrays. The cDNA libraries were enriched for flower-specific genes by subtraction with cDNA from subtending growing tissue. This strategy is meant to eliminate most household genes and numerous genes that are not specific for petals and senescence. In Iris, we spotted about 1400 clones and in carnation about 2000, of which 220 and 90 clones respectively were (partially) sequenced. Unexpectedly, during Iris senescence up-regulation was observed for many genes that previously had been characterized as being defence-related. Although such genes were also found in carnation, their relative contribution to the changes in expression seemed less pronounced. Another remarkable result was the limited number of known ethylene-related genes in carnation that were detected. Among those found was ACO1. Other ethylene-related genes may have been lost in the subtraction; and ACO1 seems specific for the ethylene climacteric. No ethylene-related genes were found in Iris. Since ethylene does not regulate petal senescence in Iris this is no surprise. Some similarities were found between Iris and carnation. In both species a considerable proportion of the up-regulated genes encode enzymes that are involved in the degradation of lipids, protein, and complex carbohydrates such as cell walls. Several genes involved in signal transduction and in transcription were observed to change expression levels in both species, but none were the same in both species, as judged from the limited sequence information. A novel EIN3 (EIL) transcription factor was discovered in carnation. The expression pattern of some putative transcription factors in carnation were expressed independently of ethylene treatment, and may be candidates for early regulators of traits such as ethylene senstivity. The detailed results on Iris have been published in the December 2003 issue of Plant Molecular Biology (53: 845-865); the results on carnation have been submitted

Multiple mediators of plant programmed cell death : interplay of conserved cell death mechanisms and plant-specific regulators

Programmed cell death (PCD) is a process aimed at the removal of redundant, misplaced, or damaged cells and it is essential to the development and maintenance of multicellular organisms. In contrast to the relatively well-described cell death pathway in animals, often referred to as apoptosis, mechanisms and regulation of plant PCD are still ill-defined. Several morphological and biochemical similarities between apoptosis and plant PCD have been described, including DNA laddering, caspase-like proteolytic activity, and cytochrome c release from mitochondria. Reactive oxygen species (ROS) have emerged as important signals in the activation of plant PCD. In addition, several plant hormones may exert their respective effects on plant PCD through the regulation of ROS accumulation. The possible plant PCD regulators discussed in this review are integrated in a model that combines plant-specific regulators with mechanisms functionally conserved between animals and plant

Cloning and analysis of a defender against apoptotic cell death (DAD1) homologue from tomato

A cDNA clone homologous to the human defender against apoptotic cell death (DAD1) gene, which is believed to be a conserved inhibitor of programmed cell death, was isolated from tomato (Lycopersicon esculentum cv. Prisca). The 351 basepairs open reading frame predicted a 116 amino acid protein sequence (LeDAD1) that showed high homology to other DAD1 proteins. Northern analysis revealed that LeDAD1 was constitutively expressed during ripening of wildtype, rin, and Nr tomato fruit

Apoptotic-like cell death marks the early stages of gypsophila (Gypsophila paniculata) petal senescence

Flower senescence is accompanied by the rapid death of large numbers of cells. In situ DNA degradation was studied in gypsophila (Gypsophila paniculata) petals during flower opening and senescence, using terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL). In flowers that had just opened, TUNEL staining was exclusively observed in nuclei in the vicinity of (developing) vessels, presumably reflecting xylem vessel differentiation. In fully open flowers, TUNEL positive nuclei appear in all the cells well before the increase in ethylene production and visible signs of senescence. Occurrence of TUNEL positive nuclei was stimulated by ethylene but not prevented by silver thiosulphate. The results show that gypsophila petal senescence is a form of PCD with features of animal apoptosis and suggest that nuclear DNA degradation is an early regulatory event rather than a result of massive cell death in the final stage of senescence

Ethylene perception is required for the expression of tomato ripening-related genes and associated physiological changes even at advanced stages of ripening

Treatment of tomato fruit (Lycopersicon esculentum L. cv Prisca) with 1-methylcyclopropene (1-MCP), a potent inhibitor of ethylene action, delayed colour development, softening, and ethylene production in tomato fruit harvested at the mature green breaker, and orange stages. 1-MCP treatment also decreased the mRNA abundance of phytoene synthase 1 (PSY1), expansin 1 (EXP1), and 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase 1 (ACO1), three ripening-related tomato genes, in mature green, breaker, orange, and red ripe fruit. These results demonstrate that the ripening process can be inhibited both on a physiological and molecular level, even at very advanced stages of ripening. The effects of 1-MCP on ripening lasted 5–7 days and could be prolonged by renewed exposure

A tomato metacaspase gene is upregulated during programmed cell death in Botrytis cinerea-infected leaves

Programmed cell death (PCD) in plant cells is often accompanied by biochemical and morphological hallmarks similar to those of animal apoptosis. However, orthologs of animal caspases, cysteinyl aspartate-specific proteases that constitute the core component of animal apoptosis, have not yet been identified in plants. Recent studies have revealed the presence of a family of genes encoding proteins with distant homology to mammalian caspases, designated metacaspases, in the Arabidopsis thaliana genome. Here, we describe the isolation of LeMCA1, a type-II metacaspase cDNA clone from tomato (Lycopersicon esculentum Mill.). BLAST analysis demonstrated that the LeMCA1 gene is located in close vicinity of several genes that have been linked with PCD. Southern analysis indicated the existence of at least one more metacaspase in the tomato genome. LeMCA1 mRNA levels rapidly increased upon infection of tomato leaves with Botrytis cinerea, a fungal pathogen that induces cell death in several plant species. LeMCA1 was not upregulated during chemical-induced PCD in suspension-cultured tomato cells