Diversity and activity of sugar transporters in nematode-induced root syncytia

The plant-parasitic nematode Heterodera schachtii stimulates plant root cells to form syncytial feeding structures which synthesize all nutrients required for successful nematode development. Cellular re-arrangements and modified metabolism of the syncytia are accompanied by massive intra- and intercellular solute allocations. In this study the expression of all genes annotated as sugar transporters in the Arabidopsis Membrane Protein Library was investigated by Affymetrix gene chip analysis in young and fully developed syncytia compared with non-infected Arabidopsis thaliana roots. The expression of three highly up-regulated (STP12, MEX1, and GTP2) and three highly down-regulated genes (SFP1, STP7, and STP4) was analysed by quantitative RT-PCR (qRT-PCR). The most up-regulated gene (STP12) was chosen for further in-depth studies using in situ RT-PCR and a nematode development assay with a T-DNA insertion line revealing a significant reduction of male nematode development. The specific role of STP12 expression in syncytia of male juveniles compared with those of female juveniles was further shown by qRT-PCR. In order to provide evidence for sugar transporter activity across the plasma membrane of syncytia, fluorescence-labelled glucose was used and membrane potential recordings following the application of several sugars were performed. Analyses of soluble sugar pools revealed a highly specific composition in syncytia. The presented work demonstrates that sugar transporters are specifically expressed and active in syncytia, indicating a profound role in inter- and intracelluar transport processes

The EFF-1A Cytoplasmic Domain Influences Hypodermal Cell Fusions in C. elegans But Is Not Dependent on 14-3-3 Proteins.

BACKGROUND: Regulatory and biophysical mechanisms of cell-cell fusion are largely unknown despite the fundamental requirement for fused cells in eukaryotic development. Only two cellular fusogens that are not of clear recent viral origin have been identified to date, both in nematodes. One of these, EFF-1, is necessary for most cell fusions in Caenorhabditis elegans. Unregulated EFF-1 expression causes lethality due to ectopic fusion between cells not developmentally programmed to fuse, highlighting the necessity of tight fusogen regulation for proper development. Identifying factors that regulate EFF-1 and its paralog AFF-1 could lead to discovery of molecular mechanisms that control cell fusion upstream of the action of a membrane fusogen. Bioinformatic analysis of the EFF-1A isoform\u27s predicted cytoplasmic domain (endodomain) previously revealed two motifs that have high probabilities of interacting with 14-3-3 proteins when phosphorylated. Mutation of predicted phosphorylation sites within these motifs caused measurable loss of eff-1 gene function in cell fusion in vivo. Moreover, a human 14-3-3 isoform bound to EFF-1::GFP in vitro. We hypothesized that the two 14-3-3 proteins in C. elegans, PAR-5 and FTT-2, may regulate either localization or fusion-inducing activity of EFF-1. METHODOLOGY/PRINCIPAL FINDINGS: Timing of fusion events was slightly but significantly delayed in animals unable to produce full-length EFF-1A. Yet, mutagenesis and live imaging showed that phosphoserines in putative 14-3-3 binding sites are not essential for EFF-1::GFP accumulation at the membrane contact between fusion partner cells. Moreover, although the EFF-1A endodomain was required for normal rates of eff-1-dependent epidermal cell fusions, reduced levels of FTT-2 and PAR-5 did not visibly affect the function of wild-type EFF-1 in the hypodermis. CONCLUSIONS/SIGNIFICANCE: Deletion of the EFF-1A endodomain noticeably affects the timing of hypodermal cell fusions in vivo. However, prohibiting phosphorylation of candidate 14-3-3-binding sites does not impact localization of the fusogen. Hypodermal membrane fusion activity persists when 14-3-3 expression levels are reduced

The anatomical structure of the common fir (Abies alba Mili.) bark. I. Development of bark tissues

Qualitative and quantitative changes in bark tissues of the fir (Abies alba) connected with the long-term cambium activity were investigated: 1) the development of secondary phloem elements forming the annual ring in different-aged parts of the stem was studied in detail; 2) the terms of formation of the phloem elements were established as well as the duration of the periods of their differentiation and functioning; 3) observations were made of the changes occurring in nonconducting phloem and primary cortex tissues; 4) the course of development of the periderm was studied

The anatomical structure of the common fir (Abies alba Mill.) bark. II. Quantitative changes in bark tissues within the stem

The regularities of the process of secondary sclerification of secondary phloem were investigated. A correlation was found between the number of sclereid layers and the age of the tree. A new sclereid layer forms within the secondary phloem on the average every 4-8 years. The process of formation of the successive layers is not uniform in its course. The differences concern mainly the structure of the successive sclereid layers and the duration of the process of their filling

Rozwój warstw ściennych i otwieranie się pylnika u kilku dzikich gatunków ziemniaka (Solarium chacoense Bitt., S. phureja Juz. et Buk., 5. giberulosum Juz. et Buk., S. Commersonii Dun.) [The development of the parietal layers and dehiscence of the anthers at some wild potato species (Solanum chacoense Bitt., S. phureja Juz. et Buk., S. giberulosum Juz. et Buk., and S. Commersonii Dun.]

The anthers in these species like in whole genus Solanum are poricidal still some differences have been observed in structure of their anther walls. The fibrous cells associated with dehiscence restricted usually to the area around the pore may form a single layer or may form compact mass often irregular in thickness or it may be lacking at all. Besides the apical pore one can observe a lateral dehiscence. By desintegra-tion of cells a gap is forming in the wall separating the two anther chambers which causes breaking it down. Then stomium like cells in epidermis contribute to the lateral opening

The anatomical structure of the common fir (Abies alba Mili.) bark. I. Development of bark tissues

Qualitative and quantitative changes in bark tissues of the fir (Abies alba) connected with the long-term cambium activity were investigated: 1) the development of secondary phloem elements forming the annual ring in different-aged parts of the stem was studied in detail; 2) the terms of formation of the phloem elements were established as well as the duration of the periods of their differentiation and functioning; 3) observations were made of the changes occurring in nonconducting phloem and primary cortex tissues; 4) the course of development of the periderm was studied

The reorganization of root anatomy and ultrastructure of syncytial cells in tomato [Lycopersicon esculentum Mill.] infected with potato cyst nematode [Globodera rostochiensis Woll.]

The sequence of anatomical and ultrastructural events leading to the syncytium development in tomato roots infected with Globodera rostochiensis was examined. The syncytia were preferentially induced in cortical or pericyclic cells in the elongation zone of root. They developed towards the vascular cylinder by incorporation of new cells via local cell wall breakdown. After surrounding primary phloem bundle and reaching xylem tracheary elements syncytia spread along vascular cylinder. Roots in primary state of growth seemed to be the best place for syncytium induction as syncytia formed in the zone of secondary growth were less hypertrophied. At the ultrastructural level syncytial elements were characterized by strong hypertrophy, breakdown of central vacuole, increased volume of cytoplasm, proliferation of organelles, and enlargement of nuclei. On the syncytial wall adjoining vessels the cell wall ingrowths were formed, while the syncytial walls at interface of phloem were considerably thickened. They lacked of functional plasmodesmata and did not form any ingrowths. Using immunofluorescent-labelling and immunogold-labelling methods tomato expansin 5 protein was localized in nematode infected roots. The distribution of LeEXP A5 was restricted only to the walls of syncytia. The protein distribution pattern indicated that LeEXP A5 could mediates cell wall expansion during hypertrophy of syncytial elements

Immunolocalization of alfa-expansin protein (NtEXPA5) in tobacco roots in the presence of the arbuscular mycorrhizal fungus Glomus mosseae Nicol. and Gerd.

The arbuscules of mycorrhizae develop within apoplastic compartments of the host plant, as they are separated from the cell protoplast by an interfacial matrix continuous with the plant cell wall. Expansins are proteins that allow cell wall loosening and extension. Using fluorescence and electron microscopy we located the NtEXPA5 epitopes recognized by polyclonal antibody anti-NtEXPA5 in mycorrhizal tobacco roots. The expansin protein was localized mainly within the interfacial matrix of intracellular hyphae, arbuscule trunk and main branches. NtEXPA5 proteins were detected neither within the interface of collapsing arbuscule branches nor in non-colonized cortex cells. In plant cell walls, expansin protein was detected only at the penetration point and in the parts of cell walls that adhered firmly to fungal hyphae growing intracellularly. For the first time, NtEXPA5 protein was localized ultrastructurally in hyphae growing intracellularly at the interface of the hypha tip and sites of bending. The novel localization of NtEXPA5 protein suggests that this protein may be involved in the process of arbuscule formation: that is, in promoting apical hyphal growth and arbuscule ramification, as well as in controlling the dynamic of arbuscule mycorrhiza development