Ultrastructure Studies And Subcellular Localization Of Allantoicase In Dictyostelium Discoideum During Cell Differentiation And Apoptosis

Dictyostelium discoideum, commonly referred to as a protozoan, is a unicellular eukaryote that starts its life as a unicellular amoeba, and multiplies by binary fission. Bacteria and yeast are the main source of food of the D. discoideum. Once the food becomes sparse they aggregate to form a multicellular fruiting body composed of two main cell types:stalk cells that support a spore-containing sorus. Spores are protected by a tough cell wall. The stalk raises the spore head high enough for the spores to be scattered away to maximize the possibility of germination in a more favorable environment. As the formation of these multicellular stages, are homologous to embryonic development in higher eukaryotes, include physiological and biochemical mechanisms such as cell-cell adhesion, cell differentiation and apoptosis, D. discoideum is an important model organism for cytobiology research.Although a preliminary knowledge of the morphogenesis of multi cellular development was gained in the previous research, the exact ultrastructural characteristics inside the cells, and the metabolic function of the organelles remain unknown. By means of transmission electron microscopy, the ultrastructure intracellular of D. discoideum during unicellular growth was observed. The ultrastructure characterisitics of the cells indicated all metabolism-related organelles were well-developed, this phenomenon illustrates that the unicellular amoeba conducts exuberant physiological metabolic activities, and the morphological characteristics of the organelles can offer reference for further study of the ultrastructure mechanism and the roles of the Signal Proteins during multi cellular development.Using this polyclonal antibody and colloidal gold as labels, immunoelectron microscopy technique was adopted to analyze the changes of intracellular allantoicase distribution in D. discoideum cells during log phase, aggregation stage, mound stage, slug body stage and fruiting stage. The analysis of the alllantoicase subcellular location and morphological features of organelles showed, in logarithmic phase allantoicase was mainly distributed in food vacuoles, since earlier research suggested that bacteria may express alllantoicase, the bacteria in vacuoles may be the origin of these allantoicase; during aggregation stage this protein was mainly observe in the mitochondria and the cytoplasm surrounding them, which supports Hayashi's research conclusions for amphibians; At mound stage, the allantoicase inside the cells which will form pre-spore cells, was located in the vacuoles inside the mitochondria undergoing endoautophagic apoptosis. As the pre-spore cells differentiating into resting cell, the allantoicase decreased and disappeared in those unbroken mitochondria.While almost all Organelles in dying pre-stalk cells contained allantoicase, until stalk cell formed. This result show that the two types of cells which will differentiate into spore cells and stalk cells, have apoptosis-related intracellular location of allantoicase, and suggests the allantoicase involves the regulation of cell differentiation and apoptosis.