The development of multicellular organisms involves an important balance between cell growth, cell division and cell death. In animals, programmed cell death (PCD) plays a key role by forming and... Show moreThe development of multicellular organisms involves an important balance between cell growth, cell division and cell death. In animals, programmed cell death (PCD) plays a key role by forming and deleting structures, controlling cell numbers and eliminating abnormal damaged cells. Caspases were found to be the key executioner of the cell suicide pathway. In plants, PCD plays an important role in development and in the responses to pathogens and abiotic stresses. Some common features of PCD were found to be conserved in both plants and animals (cytoplasm shrinkage, cytochrome c leakage out of mitochondria, chromatin condensation, altered nuclear morphology, DNA fragmentation in large fragments and DNA laddering). This indicates that a similar apoptotic machinery may be present in plants. After the elucidation of the complete sequences of Arabidopsis and rice, it has become clear that no genes for the caspases are present in plants. In this thesis, three model systems are described to determine the existence of caspase-like activities in plants during PCD. The first model used is a T-DNA integration arabidopsis mutant that shows necrotic spots on its leaves. The second model used is barley, during the transition from multicellular structures to globular embryos during androgenesis. The third model used was an artificial model of rice suspension cells with heat-shock as a PCD inducer. In these models, PCD was described and caspase-like activity was measured. In addition, the development and implementation of the protocol to purify plant caspase-3 like activity is described. Show less
Maraschin, S. de F.; Gaussand, G.M.D.J.; Pulido, A.; Olmedilla, A.; Lamers, G.E.; Korthout, H.A.; ... ; Wang, M. 2005
Androgenesis represents one of the most fascinating examples of cell differentiation in plants. In barley, the conversion of stressed uninucleate microspores into embryo-like structures is highly... Show moreAndrogenesis represents one of the most fascinating examples of cell differentiation in plants. In barley, the conversion of stressed uninucleate microspores into embryo-like structures is highly efficient. One of the bottlenecks in this process is the successful release of embryo-like structures out of the exine wall of microspores. In the present work, morphological and biochemical studies were performed during the transition from multicellular structures to globular embryos. Exine wall rupture and subsequent globular embryo formation were observed only in microspores that divided asymmetrically. Independent divisions of the generative and the vegetative nuclei gave rise to heterogeneous multicellular structures, which were composed of two different cellular domains: small cells with condensed chromatin structure and large cells with normal chromatin structure. During exine wall rupture, the small cells died and their death marked the site of exine wall rupture. Cell death in the small cell domain showed typical features of plant programmed cell death. Chromatin condensation and DNA degradation preceded cell detachment and cytoplasm dismantling, a process that was characterized by the formation of vesicles and vacuoles that contained cytoplasmic material. This morphotype of programmed cell death was accompanied by an increase in the activity of caspase-3-like proteases. The orchestration of such a death program culminated in the elimination of the small generative domain, and further embryogenesis was carried out by the large vegetative domain. To date, this is the first report to show evidence that programmed cell death takes part in the development of microspore-derived embryos. Show less
