Yu. V. Sidorchuk

Characteristics of the cytomictic channel formation in Nicotiana tabacum L. pollen mother cells

Electron-microscopic analysis of cytomictic channels formation in the pollen mother cells in tobacco at the stage of meiosis prophase I of anthers has been conducted. The cytomictic channels in the pollen mother cells in tobacco have been established to be formed under the basis of both single plasmodesmata and de novo with the involvement of specific electron-dense bodies. The role of cytomictic channels in microsporogenesis regulation is discussed.

Peculiarities of cytomixis in pollen mother cells of transgenic tobacco plants (Nicotiana tabacum L.) with mutant phenotype

The frequency characteristics and cytological picture of cytomixis in the course of male meiosis are described in transgenic tobacco plants (Nicotiana tabacum L.) with altered flower morphology and male sterility. Effects of cytomixis on qualitative composition of meiotic products are studied (formation of cytoplasts and polyads). Doubling of the chromosome number was established to increase frequency of cytomixis in the studied plants.

Role of Microtubular Cytoskeleton and Callose Walls in the Manifestation of Cytomixis in Pollen Mother Cells of Tobacco Nicotiana tabacum L.

The structure and dynamics of microtubular cytoskeleton and of callose walls in normal pollen mother cells (PMC) of tobacco N. tabacum L. and in cells with intercellular translocation of nuclear material (cytomictic) was studied in the course of the cell cycle. The microtubular cytoskeleton was established as playing no obvious role in the process of cytomixis. The elevated level of cytomictic seems to be due to disturbances of synthesis of callose walls as a result of their attenuation and perforation. Possible causes of cytomictic in tobacco PMC at the cellular level are discussed.

Comparative analysis of HBV M-antigen production in leaves of individual transgenic carrot plants

76 The advantages of plants as alternative systems for expression of heterologous proteins are illustrated by numerous examples of accumulation in plant tissues of complex functionally active immunoglobulins, secretory granules, collagen, hemoglobin, and cytokines [1]. Transgenic plants accumulating antigens against various pathogens can be used as edible vaccines, including those against hepatitis B virus [2].

Premature Cytokinesis in Pollen Mother Cells of Transgenic Tobacco Plants ( Nicotiana tabacum L.)

For the majority of dicotyledonous plants, cytokinesis in PMC is staged only once, i.e., after the completion of two cycles of caryokinesis. In the article, a cytological picture and the frequency characteristics of anomalies are shown, in which the cytokinesis in the PMCs of transgenic tobacco plants was already initiated after the first meiotic division. It is shown that, in such cells, the basic processes of cytoskeletal reorganization, which is typical for the simultaneous type of cytokinesis, remained unmodified. However, in most cases, premature division of cytoplasm took place with abnormalities, e.g., with the formation of a membranous “tunnel” or “gash.” It has been detected that the initialization of an additional round of cytokinesis is not an obstacle to the performance of the nuclear cycle or cytokinesis after the second meiotic division. Thus, in the presence of this anomaly, there is a change in the type of cytoplasm division, i.e., of simultaneous to successive.

Cytomixis in Mother Pollen Cells of Transgenic Tobacco (Nicotiana tabacum L.) Plants

Cytomixis—the migration of nuclei and nuclear material from one mother pollen cell to another—is widespread in nature; it has been described for many plant species [1–4]. In most cases, cytomixis was observed in mother pollen cells [2, 5–10]. Nuclei and chromosomes migrate along cytomictic channels, which join anther meiocytes in a united system [4]. It was shown that, in some plant species, cytomixis is expressed only in the prophase of the first meiotic division [2, 7, 8, 10]. In other species, chromosomal migration may last to the stage of tetrads [5, 6, 9, 11]. It was assumed that this phenomenon is of evolutional importance, because it leads to aneuploidy and polyploidy of gametes [7, 9, 10] and may be the source of B-chromosomes [5]. The researchers have not still reached a consensus of opinion on factors that cause the extrusion of nuclear material and the mechanisms that underlie this phenomenon. It is believed that cytomixis may caused by various disturbances in the structure of cytomictic channels [1, 4], synthesis of callose coat [5, 9], cell wall formation during premeiotic mitoses [6], and structure of the radial cytoskeleton [8]. Some authors believe that this phenomenon is characteristic of the plants that exhibit genetic instability (hybrids and mutants) [1, 4, 7, 10, 11].

Deformation of nuclei and abnormal spindles assembly in the second male meiosis of polyploid tobacco plants

The bipolar spindle is a major cytoskeletal structure, which ensures an equal chromosome distribution between the daughter nuclei. The spindle formation in animal cells depends on centrosomes activity. In flowering plant cells the centrosomes have not been identified as definite structures. The absence of these structures suggests that plants assemble their spindle via novel mechanisms. Nonetheless, the cellular and molecular mechanisms controlling the cytoskeleton remodeling during the spindle development in plants are still insufficiently clear. This article describes the results of a comparative analysis of the microtubular cytoskeleton dynamics during assembly of the second division spindle in tobacco microsporocytes with the normal and deformed nuclei. According to our observations, the bipolar spindle fibres are formed from short arrays of the disintegrated perinuclear cytoskeleton system, the perinuclear microtubular band. The microsporocytes of polyploid tobacco plants with deformed nuclei entirely lack this cytoskeleton structure. In such type of cells the overall prometaphase events are blocked, and the assembly of second division spindles is completely arrested.

Intra- and intertissular cytomictic interactions in the microsporogenesis of mono- and dicotyledonous plants

Comparative cytological analysis of intra- and intertissular cytomictic interactions in the microsporogenesis of mono- and dicotyledonous plants has been performed for two cellular systems: the microsporocytes and the tapetum. Cytomixis was shown to be more common for intratissular interactions, and cytomixis in the tapetum exhibited taxon-specific features, both structural and temporal. Nuclear migration in the microsporocytes mostly occurred during the zygotene–pachytene and exhibited certain synchrony with cytomixis in the tapetum. Intertissular cytomictic interactions (between the tapetum and the microsporocytes) were detected only in monocotyledonous plant anthers. Intertissular interactions may reflect more intense competition for space between the tapetum and the microsporocytes during the differentiation of anther tissues. The polyploid nuclei of the tapetum and the syncytia are powerful acceptors that can compete with the microsporocytes and attract the chromatin during translocation of the latter. The absence of intertissular interactions in dicotyledonous plants may be indicative of a better balance between the processes of differentiation of somatic and generative tissues of the microsporangium as compared to monocotyledonous plants.

Migration of DNA-Containing Organelles between Tobacco Microsporocytes during Cytomixis

Ultrastructural analysis of intercellular migration of DNA-containing organelles (nuclei, mitochondria, and plastids) in tobacco microsporogenesis during cytomixis was conducted. It was demonstrated for the first time that the migrating part of the nucleus is covered with ribosomes and can contain the accumulation of nuclear pores. The possibility of mitochondrial migration between the plant cells was proven for the first time. It was demonstrated that mitochondria extremely rarely pass into neighboring cells, and their movement occurs through one cytomictic channel. In turn, plastids can generate the accumulations around cytomictic channels and actively migrate between the cells, even through small size cytomictic channels. It was established that plastids can pass into another cell through one or several cytomictic channels, and several plastids can also simultaneously migrate through one channel. The consequences of migration of DNA-containing organelles in the cells producing the pollen are discussed.

Simple and reliable system for transient gene expression for the characteristic signal sequences and the estimation of the localization of target protein in plant cell

The efficiency of transient gene expression in plants credibly demonstrated characteristics of gene functions in numerous studies. Two key strategies of transient expression became favorites among researchers: protoplast transfection and agroinfiltration. Each of them, alongside the advantages, has its own constraints. In this work, an easy, rapid, and reliable system for characterization of the signal sequences and determinations of target protein localization in a plant cell is proposed and tested. This system—called the AgI–PrI—implies production of protoplasts from plant tissues after agroinfiltration. Reliability of the proposed system for transient gene expression has been proved using characterized signal sequences in Nicotiana benthamiana cells. The corresponding protocol is less expensive and depends to a lesser degree on the professional skills in the area of protoplast isolation and transfection; furthermore, it may be applicable to other plant species with either available efficient methods of agroinfiltration and protoplast isolation or with the potential for one of the protocols to be supplemented. Thus, the AgI–PrI technique makes it possible to combine the advantages of two widely used methods for the transient gene expression in plants—agroinfiltration and protoplast isolation and transfection—and concurrently avoids their critical points.