Programmed Cell Death Involved In The Developmental Process Of Several Plant Secretory Structures

Plant secretory structures are important structural and chemical defence system for plant protecting themselves against herbivores, pathogenic microbes, parasite and mechanical wounding and so on. Secretory structures of plant have long been studied, and most of viewpoints are educed from histological points of role or by investigation of mature secretory structures. However, studies on cytological essence of plant secretory structures such as origin, formation and senescence process are limit, and a detailed study on cytological changes is still lacking during secretory structure development. In the present work, cytological chemistry, light microscopy, scanning electron microscopy and tranmission electron microscopy were used to determine the morphology, distribution and ontogeny of secretory canals in fruits of Decaisnea fargesii Franch. (Lardizabalaceae), pigment glands in leaves of Gossypium hirsutum L. (Malvaceae), and secretory structures and non-glandular hairs of Dictamnus dasycarpus Turcz. (Rutaceae), and reveal cytological characters duiring PCD of glands and non-glandular hairs. Ultimately, an explaination on development and evolution of these glands and non-glandular hairs are also determined.Decaisnea fargesii is a latex-producing plant that is characterized by the presence of a special type of secretory structure in the pericarp, the laticiferous canal, in which the rubber granules occur and accumulate. The secretory canal is derived from epidermal cells of the fruit through a process of invagination, closing, and lysigenous activity, forming a reticulate intercellular canal system. During the mature stage, the cell walls of the secretory epidermal cells become noticably thinner and even partly lyse. The laticiferous canal is essentially a special secretory canal formed from one layer of epidermal cells, distinguished by partial cell wall lysis, with numerous secretory epidermal cells filled with abundant rubber in mature.Several hallmarks reveal forms of autophagy, indicating that formation of laticiferous canal in fruits of Decaisnea fargesii is one type of developmentally regulated PCD phenomenon. The results indicate that several morphological hallmarks such as collapse of vacuole and plasma membrane, membranous cell wall, part of cell wall degradation, disorganization of cytoplasm, the presence of numerous membrane structures and diffused flocculent material coupled with plastids, mitochondria and nucleus in the vacuole, the deformed nucleus is observed in vacuole or enveloped by rubber granules. The nuclei of the secretory epidermal cells become TUNEL positive from the sunken stage to the late expanding stage, then DAPI-negative during the mature stage, indicating an early event of DNA cleavage and a late event of complete DNA degeneration. Gel electrophoresis indicates that DNA cleavage is random and does not result in the laddering pattern indicating multiples of internucleosomal units.During the PCD of secretory epidermal cells, the rubber granules continue to be synthesized and accumulated in the secretory epidermal cells. Formation of rubber granules is random, and the precursors of rubber granules are characterized by the osmiophilic flocculent material or membranous mass structures which are close to degenerated plastids, mitochondria, or on the end of swelling endoplasmic reticulums. Later, degenerated cellular component continually integrate into the rubber precursors, and the volume of the rubber granules expands sequentially. The rubber precursors or rubber granules are then accumulated within the central vacuole by autophagy, and integrate with the vesicular, flocculent and mass structures continually, even the organelle enveloped by the rubber granule was also observed, finally shapely mature rubber granules form. During the PCD process of laticiferous canal of Decaisnea fargesii, components from cellular disintegration are used to synthesize rubber granules.Cotton is the most important industrial crop which is widely used all over the world. The genus Gossypium and its relative species in Malvaceae are characterized by pigment glands which play important roles in cotton developmental process. The pigment glands originate from clusters of initial cells distinguishable from other cells in ground meristem by their smaller sizes, dense cytoplasm, large nuclei, small vacuoles, and thin cell walls. During the developmental process, the internal cells and part of sheath cells lyse. At maturity, the pigment glands or secretory cavities consist of a wide lumen surrounded by one layer of secretory cells and 1 to 3 layers of sheath cells that became very flat and degenerated, and the outer 1-2 layer of sheath cells are evidently modified by thick cell walls.The result indicates that lysigenous formation of pigment glands is also a typical example of developmentally regulated PCD. Nuclei of the internal cells in the pigment gland-forming tissue are TUNEL-positive, and DAPI staining shown that misshapen of nuclei later degenerated, smeared bands and a lack of laddering after gel electrophoresis indicate that DNA cleavage is random, these of which are similer to nuclei and genomic DNA degeneration of the laticiferous canal of Decaisnea fargesii. Degeneration of pigment glands are distinguished by disintegration of internal cells, condensed chromosome and dense nuclei, completely degeneration of nuclei, plenty of electron-dense bodies and membrane-bound spherical structures, swollen cell walls completely collapse later, indicating on-going autophagy and autolysis.The PCD process of pigment glands in Gossypium hirsutum couples with gossypol synthesis and accumulation. During the earliest developmental stage of pigment glands, no black gossypol deposition is observed. Later, gossypol within cytoplasm and on the surface of autophagosomes increase and become larger, which process is accompanied cytoplasm and organelle degeneration, and no gossypol deposition is observed in negative group. However rarely a direct evidence about gossypol occurred in plastids or endoplasmic reticulum were also observed. In all, gossypol occurs in cytoplasm at random, pigment glands are important site for gossypol accumulation. In vacuole, the material enveloped by autophagosomes develops into black gossyple which is released after disintegration of autophagosomes and secretory and sheath cells. The results indicate that degeneration material from pigment glands PCD may be used to synthesize gossypol, gossypol synthesizing is correlated with autophagosomes, and vacuole-mediated autophagy is hypothesized to be one of the important modes for gossypol synthesis.Dictamnus dasycarpus is characterized by the presence of capitate glandular hair, trichome cavity, secretory cavity and one type of non-glandular hair. The secretory cavities were distributed only within the leaf, mainly close to adaxial epidermal cells. Mature secretory cavities consist of sheath cells, one layer of flattened secretory epidermis and a large cavity within the center. The capitate glandular hairs and the trichome-like secretory cavities were situated on the surface of leaf, floral axis, sepal, petal, filament and ovary. The capitate glandular hairs were characterized by a spherical multicellular head, two line stalk cells and two basal cells. The trichome-like cavity shared a similar shape with glandular hairs by an obvious multicellular head and a short stalk, non-glandular hairs with a single beak-shaped apex, and a sub-epidermal secretory cavity delimited by one or two layers of cells within the head. Non-glandular hair is unicellular hair which distribution is similar with glandular hairs, however no non-glandular hair is observed on filament.Non-glandular hairs, capitate glandular hairs and secretory cavities of Dictamnus dasycarpus are entirely initiated from single protoepidermal cell. Non-glandular hairs and capitate glandular hairs develop directly from a single protoepidermal cell. The secretory cavities and trichome-like cavities both originate from a single, initial epidermal cell that undergoes a periclinal division, which forms two sister cells, and the inner cell develops towards outside and inside to form trichome-like cavities and secretory cavities. Formation of trichome cavities and secretory cavities are lysigenous for collapse of internal cells or secretory cells during the developmental process. Histochemical studies by Sudan black B suggests that the trichome cavities, capitate glandular hair and secretory cavity glands are important sites for essential oils.Ultrastructural analysis shows that the disorganization of the cytoplasm is accompanied by misshapen nuclei with condensed chromatin, plasmolysis, and disintegration of the plasma membrane system followed by autolysis, where the mitochondria and degenerated plastids with disorganized membrane systems are engulfed by vacuoles, multivesicular bodies, and double-membranous autophagosomes within the vacuoles. A strong structural twist and swelling of the internal cell walls ultimately leads to complete breakdown. Nuclei of the inner internal cells within the trichome-like cavity become TUNEL-positive and DAPI-negative first; later this is detected in the outer internal cells, indicating a centrifugal nuclear degradation process. On the basis of this work, it is interesting to speculate that the lysigenous formation of the trichome-like cavity is a typical programmed cell death process. Light microscope and ultrastructural analysis also show that non-glandular hairs, capitate glandular hairs and secretory cavities of Dictamnus dasycarpus are also characterized by PCD characters, including disorganization of membrane system, double-membranous structures, multivesicular bodies, plenty of flocculent meterial and degeneration of nuclei and so on.In conclusion, although different develomental processes occur in these secretory structures, the ultimate PCD fate of these secretory structures is similar, and the secretory materials synthesize during the PCD process. It is hypothesized that PCD may play important roles in gland development of plants, and PCD's function is used for synthesizing secretions.