Marieluise Weidinger

Ca2+-requirement for a blue-light-induced chloroplast translocation inEremosphaera viridis

SummaryThe light-mediated translocation of chloroplasts inEremosphaera viridis is dependent on blue light near 450 nm, while wavelengths longer than 500 nm are inactive. The plastid translocation results in an aggregation of the organelles close to the nucleus in the center of the cell. After cessation of irradiation, the cells begin to redistribute their plastids in the cytoplasm immediately. Treatments that alter the Ca2+ concentration in the cytoplasm ofEremosphaera suggest that the translocation is regulated by calcium. Ultrastructural investigation ofEremosphaera reveals a very characteristic, multilayered and highly-ordered cell wall.

Ultrastructure of five Euglena species positioned in the subdivision Serpentes

Within the genus Euglena, the subgroup “Serpentes” is characterised by species with long, slim cell bodies, which move without flagellum by snake-like locomotion in the detritus or in the mud, or swim freely in the water with a flagellum. Two major groups can be distinguished. The first is centred around the species Euglena satelles, with Euglena carterae, Euglena adhaerens and others, and is characterised by a straight-ended anterior part of the cell without a protruding flagellum. The second group is centred around the species Euglena deses, with its varieties, and Euglena ehrenbergii, and is characterised by a lateral canal opening at the anterior end with one flagellum protruding sideways. The representatives of the whole Serpentes group have various (15–30) large chloroplasts containing characteristic naked pyrenoids. The exception is Euglena ehrenbergii, which possesses innumerable small chloroplasts without pyrenoids. To better characterise this whole subgroup, to better taxonomically distinguish between the diverse species and to provide a basis for further molecular-genetic analysis of the phylogeny of and relationship between the Euglena species, we used transmission and scanning electron microscopy to investigate the five selected species. One important distinguishing feature among the species is the form of the pellicle. It can differ in thickness or cross-sectional shape (e.g. A-, M-or plateau-like shape) and can have various arrangements of microtubules and endoplasmic reticulum mucus vesicles. We show that the group is more heterogeneous than expected and that some species have very individual features that poorly fit into a common Serpentes group, particularly the above-mentioned Euglena ehrenbergii. Euglena carterae, formerly named Euglena deses var. carterae, with its typical straight-ended canal opening, does not fit into the Euglena deses varieties, as has already been confirmed by molecular genetic methods.

Brefeldin A induces callose formation in onion inner epidermal cells.

SummaryThe antibiotic fungal toxin brefeldin A (BFA) causes synthesis of additional cell wall material in adult differentiated onion inner epidermal cells at concentrations of 5–30 μg/ml. This tertiary wall contains callose and is layered on the secondary cellulosic wall in a time- and dose-dependent manner. Initially, callose is found in pit fields in the form of small vesicular patches. With time and dose, depositions grow in size and form large plugs invaginating into the cell, where the adjacent cytoplasm forms bulky accumulations and contains many organelles including endomembranes. Within the cytoplasm, BFA exerts the characteristic morphological effects on the secretory system including changes of the Golgi stacks, formation of large vesicles, and proliferation of dilated cisternae of the endoplasmic reticulum. Higher concentrations of BFA (60 μg/ml) lead to disintegration of the Golgi apparatus; they have no effects on the cell wall, no callose synthesis occurs. We conclude from these observations that BFA has two independent targets in onion cells. BFA acts on the plasma membrane, hence operating as an elicitor of plant defense reactions and thus activates callose synthesis. BFA acts also on the membranes of the secretory system and influences budding and fusion of vesicles at the endoplasmic reticulum and at the dictyosomes. These two mechanisms occur in parallel, suggesting that the secretory system still can play its presumed role in callose synthesis. Only when dictyosomes are completely disintegrated, no more callose is formed.

Metalloid Contaminated Microhabitats and their Biodiversity at a Former Antimony Mining Site in Schlaining, Austria

This paper is on the biological impact of arsenic and antimony on the flora and microflora on a former Sb- mining site in Schlaining (Stadtschlaining, Burgenland, Austria). Several habitats were investigated with respect to biodi- versity and metalloid contamination in soil. Although the overburden of the mining activity had been remediated less than ten years ago, metalloid concentrations occurred in soil up to 1.4‰ As and 3.6% Sb, respectively, in some microhabitats, as determined by Instrumental Neutron Activation Analysis. These metalloids were embedded into a nonuniform minera- logical background. Metalloid mobility could not be explained by common models, indicating that predictions on the mo- bility of geogenic metalloids require additional mineralogical data. The biological effects of this contamination were vari- able. We observed that metalloid resistant strands of microorganisms appeared in the contaminated soil. In cultivation ex- periments, Sb was found to be more toxic than As. Sulphur oxidising strand were more resistant than organotrophic ones and grew even better on cultivation media spiked with 10 ppm As than on the unspiked control. The flora was only par- tially influenced: the lowest biodiversity was found in metalloid richest soils, but moderate contamination resulted in en- hanced species numbers. Only in one case, where the pH-buffering capacity of the soil was exceeded by consumption of the entire carbonate, no embryophytes occurred. This was probably due to extreme pH conditions as well as to metalloid concentrations. Our data support the hypothesis that higher plants are rather affected by extreme soil conditions, which of- ten coincide with As contaminations, than by the contamination itself. A small rivulet in this area contained 26 � g/l and thus exceeded the WHO guideline value for As in drinking water by a factor of 2.6. Indeed we observed a diminished bio- diversity in this rivulet.

The inhibition of systrophe in different organisms

SummaryThe phenomenon of systrophe following different stimuli was comparatively investigated in the cells of different plant organisms. In all cases systrophe could be inhibited by metabolic inhibitors. The influence of colchicine, cytochalasin B, vinblastine and APM (amiprophos-methyl) varied.

Using RT-qPCR, Proteomics, and Microscopy to Unravel the Spatio-Temporal Expression and Subcellular Localization of Hordoindolines Across Development in Barley Endosperm

Hordeum vulgare (barley) hordoindolines (HINs), HINa, HINb1, and HINb2, are orthologous proteins of wheat puroindolines (PINs) that are small, basic, cysteine-rich seed-specific proteins and responsible for grain hardness. Grain hardness is, next to its protein content, a major quality trait. In barley, HINb is most highly expressed in the mid-stage developed endosperm and is associated with both major endosperm texture and grain hardness. However, data required to understand the spatio-temporal dynamics of HIN transcripts and HIN protein regulation during grain filling processes are missing. Using reverse transcription quantitative PCR (RT-qPCR) and proteomics, we analyzed HIN transcript and HIN protein abundance from whole seeds (WSs) at four [6 days after pollination (dap), 10, 12, and ≥20 dap] as well as from aleurone, subaleurone, and starchy endosperm at two (12 and ≥20 dap) developmental stages. At the WS level, results from RT-qPCR, proteomics, and western blot showed a continuous increase of HIN transcript and HIN protein abundance across these four developmental stages. Miroscopic studies revealed HIN localization mainly at the vacuolar membrane in the aleurone, at protein bodies (PBs) in subaleurone and at the periphery of starch granules in the starchy endosperm. Laser microdissetion (LMD) proteomic analyses identified HINb2 as the most prominent HIN protein in starchy endosperm at ≥20 dap. Additionally, our quantification data revealed a poor correlation between transcript and protein levels of HINs in subaleurone during development. Here, we correlated data achieved by RT-qPCR, proteomics, and microscopy that reveal different expression and localization pattern of HINs in each layer during barley endosperm development. This indicates a contribution of each tissue to the regulation of HINs during grain filling. The effect of the high protein abundance of HINs in the starchy endosperm and their localization at the periphery of starch granules at late development stages at the cereal-based end-product quality is discussed. Understanding the spatio-temporal regulated HINs is essential to improve barley quality traits for high end-product quality, as hard texture of the barley grain is regulated by the ratio between HINb/HINa.

Expression of Genes for Si Uptake, Accumulation, and Correlation of Si with Other Elements in Ionome of Maize Kernel

The mineral composition of cells, tissues, and organs is decisive for the functioning of the organisms, and is at the same time an indicator for understanding of physiological processes. We measured the composition of the ionome in the different tissues of maize kernels by element microanalysis, with special emphasis on silicon (Si). We therefore also measured the expression levels of the Si transporter genes ZmLsi1, ZmLsi2 and ZmLsi6, responsible for Si uptake and accumulation. Two weeks after pollination ZmLsi1 and ZmLsi6 genes were expressed, and expression continued until the final developmental stage of the kernels, while ZmLsi2 was not expressed. These results suggest that exclusively ZmLsi1 and ZmLsi6 are responsible for Si transport in various stages of kernel development. Expression level of ZmLsi genes was consistent with Si accumulation within kernel tissues. Silicon was mainly accumulated in pericarp and embryo proper and the lowest Si content was detected in soft endosperm and the scutellum. Correlation linkages between the distribution of Si and some other elements (macroelements Mg, P, S, N, P, and Ca and microelements Cl, Zn, and Fe) were found. The relation of Si with Mg was detected in all kernel tissues. The Si linkage with other elements was rather specific and found only in certain kernel tissues of maize. These relations may have effect on nutrient uptake and accumulation.

Short-term influence of Cu, Zn, Ni and Cd excess on metabolism, ultrastructure and distribution of elements in lichen Xanthoria parietina (L.) Th. Fr.

Lichens are symbiotic organisms that are very sensitive to heavy metal pollution. However, there is little evidence of how heavy metal pollution affects the physiological status, ultrastructural changes and distribution of elements in the layers of lichen thalli. For this purpose we simulated metal pollution to lichens and studied its impact on Xanthoria parietina. Thalli were treated with the heavy metals Cu, Zn, Ni, Cd in the form of sulfates at concentrations of 100µM and 500µM during 24, 48 and 72h. Untreated lichens served as controls. We assessed the status of physiological parameters (fluorescence and integrity of chlorophyll a, content of soluble proteins and ergosterol), ultrastructural changes, especially to the photobiont, and the distribution of elements in the layers of thalli in relation to treatment with heavy metals. We found positive correlations between the content of all tested heavy metals and the physiological response. We assessed the toxicity of the selected metals as follows: Cd >= Cu >= Ni > Zn, based on the effects on the photobiont layer in the lichen thallus and physiological measurements.

The inhibition of systrophe by cytochalasin B

SummaryCytochalasin B inhibits the phenomenon of the systrophe following plasmolysis.

Microscopic and Proteomic Analysis of Dissected Developing Barley Endosperm Layers Reveals the Starchy Endosperm as Prominent Storage Tissue for ER-Derived Hordeins Alongside the Accumulation of Barley Protein Disulfide Isomerase (HvPDIL1-1)

Barley (Hordeum vulgare) is one of the major food sources for humans and forage sources for animal livestock. The average grain protein content (GPC) of barley ranges between 8 and 12%. Barley hordeins (i.e., prolamins) account for more than 50% of GPC in mature seeds and are important for both grain and flour quality. Barley endosperm is structured into three distinct cell layers: the starchy endosperm, which acts essentially as storage tissue for starch; the subaleurone, which is characterized by a high accumulation of seed storage proteins (SSPs); and the aleurone, which has a prominent role during seed germination. Prolamins accumulate in distinct, ER-derived protein bodies (PBs) and their trafficking route is spatio-temporally regulated. The protein disulfide isomerase (PDI) has been shown to be involved in PB formation. Here, we unravel the spatio-temporal proteome regulation in barley aleurone, subaleurone, and starchy endosperm for the optimization of end-product quality in barley. We used laser microdissection (LMD) for subsequent nanoLC-MS/MS proteomic analyses in two experiments: in Experiment One, we investigated the proteomes of dissected barley endosperm layers at 12 and at ≥20 days after pollination (DAP). We found a set of 10 proteins that were present in all tissues at both time points. Among these proteins, the relative protein abundance of D-hordein, B3-hordein and HvPDIL1-1 significantly increased in starchy endosperm between 12 and ≥20 DAP, identifying the starchy endosperm as putative major storage tissue. In Experiment Two, we specifically compared the starchy endosperm proteome at 6, 12, and ≥20 DAP. Whereas the relative protein abundance of D-hordein and B3-hordein increased between 6 and ≥20 DAP, HvPDIL1-1 increased between 6 and 12 DAP, but remained constant at ≥20 DAP. Microscopic observations showed that these relative protein abundance alterations were accompanied by additional localization of hordeins at the periphery of starch granules and a partial re-localization of HvPDIL1-1 from PBs to the periphery of starch granules. Our data indicate a spatio-temporal regulation of hordeins and HvPDIL1-1. These results are discussed in relation to the putative role of HvPDIL1-1 in end-product quality in barley.