Sponge-derived Ageladine A affects the in vivo fluorescence emission spectra of microalgae.

In several marine hosts of microalgae, fluorescent natural products may play an important role. While the ecological function of these compounds is not well understood, an interaction of these molecules with the photosynthesis of the symbionts has been suggested. In this study, the effect of Ageladine A (Ag A), a pH-dependent fluorophore found in sponges of the genus Agelas, on microalgal fluorescence was examined. The spectra showed an accumulation of Ag A within the cells, but with variable impacts on fluorescence. While in two Synechococcus strains, fluorescence of phycoerythrin increased significantly, the fluorescence of other Synechococcus strains was not affected. In four out of the five eukaryote species examined, chlorophyll a (Chl a) fluorescence intensity was modulated. In Tisochrysis lutea, for example, the position of the fluorescence emission maximum of Chl a was shifted. The variety of these effects of Ag A on microalgal fluorescence suggests that fluorophores derived from animals could play a crucial role in shaping the composition of marine host/symbiont systems.

Biomineralization of strontium and barium contributes to detoxification in the freshwater alga Micrasterias.

The unicellular model alga Micrasterias denticulata inhabits acid peat bogs that are highly endangered by pollutants due to their high humidity. As it was known from earlier studies that algae like Micrasterias are capable of storing barium naturally in form of BaSO4 crystals, it was interesting to experimentally investigate distribution and sequestration of barium and the chemically similar alkaline earth metal strontium. Additionally, we intended to analyze whether biomineralization by crystal formation contributes to diminution of the generally toxic effects of these minerals to physiology and structure of this alga which is closely related to higher plants. The results show that depending on the treatment differently shaped crystals are formed in BaCl2 and Cl2Sr exposed Micrasterias cells. Modern microscopic techniques such as analytical TEM by electron energy loss spectroscopy and Raman microscopy provide evidence for the chemical composition of these crystals. It is shown that barium treatment results in the formation of insoluble BaSO4 crystals that develop within distinct compartments. During strontium exposure long rod-like crystals are formed and are surrounded by membranes. Based on the Raman signature of these crystals their composition is attributed to strontium citrate. These crystals are instable and are dissolved during cell death. During strontium as well as barium treatment cell division rates and photosynthetic oxygen production decreased in dependence of the concentration, whereas cell vitality was reduced only slightly. Together with the fact that TEM analyses revealed only minor ultrastructural alterations as consequence of relatively high concentrated BaCl2 and Cl2Sr exposure, this indicates that biomineralization of Sr and Ba protects the cells from severe damage or cell death at least within a particular concentration range and time period. In the case of Sr treatment where ROS levels were found to be elevated, hallmarks for autophagy of single organelles were observed by TEM, indicating beginning degradation processes.

Structural stress responses and degradation of dictyosomes in algae analysed by TEM and FIB-SEM tomography.

Stress-induced physiological deficiencies in cells are reflected in structural, morphological and functional reactions of organelles. Although numerous investigations have focused on chloroplasts and mitochondria as main targets of different stressors in plant cells, there is insufficient information on the plant Golgi apparatus as stress sensor. By using the advantages of field emission scanning electron microscopy tomography in combination with classical ultrathin sectioning and transmission electron microscopic analyses, we provide structural evidence for common stress responses of the large and highly stable dictyosomes in the algal model system Micrasterias. Stress is induced by different metals such as manganese and lead, by starvation in 9 weeks of darkness or by inhibiting photosynthesis or glycolysis and by disturbing ionic homeostasis via KCl. For the first time a stress-induced degradation pathway of dictyosomes is described that does not follow "classical" autophagy but occurs by disintegration of cisternae into single membrane balls that seem to be finally absorbed by the endoplasmic reticulum (ER). Comparison of the morphological features that accompany dictyosomal degradation in Micrasterias to similar reactions observed during the same stress application in Nitella indicates an ubiquitous degradation process at least in algae. As the algae investigated belong to the closest relatives of higher land plants these results may also be relevant for understanding dictyosomal stress and degradation responses in the latter phylogenetic group. In addition, this study shows that two-dimensional transmission electron microscopy is insufficient for elucidating complex processes such as organelle degradation, and that information from three-dimensional reconstructions as provided by field emission scanning electron microscopy tomography is absolutely required for a comprehensive understanding of the phenomenon.

Rescue of heavy metal effects on cell physiology of the algal model system Micrasterias by divalent ions.

Recent studies have shown that metals such as copper, zinc, aluminum, cadmium, chromium, iron and lead cause severe dose-dependent disturbances in growth, morphogenesis, photosynthetic and respiratory activity as well as on ultrastructure and function of organelles in the algal model system Micrasterias denticulata (Volland et al., 2011, 2012; Andosch et al., 2012). In the present investigation we focus on amelioration of these adverse effects of cadmium, chromium and lead by supplying the cells with different antioxidants and essential micronutrients to obtain insight into metal uptake mechanisms and subcellular metal targets. This seems particularly interesting as Micrasterias is adapted to extremely low-concentrated, oligotrophic conditions in its natural bog environment. The divalent ions of iron, zinc and calcium were able to diminish the effects of the metals cadmium, chromium and lead on Micrasterias. Iron showed most ameliorating effects on cadmium and chromium in short- and long-term treatments and improved cell morphogenesis, ultrastructure, cell division rates and photosynthesis. Analytical transmission electron microscopic (TEM) methods (electron energy loss spectroscopy (EELS) and electron spectroscopic imaging (ESI)) revealed that chromium uptake was decreased when Micrasterias cells were pre-treated with iron, which resulted in no longer detectable intracellular chromium accumulations. Zinc rescued the detrimental effects of chromium on net-photosynthesis, respiration rates and electron transport in PS II. Calcium and gadolinium were able to almost completely compensate the inhibiting effects of lead and cadmium on cell morphogenesis after mitosis, respectively. These results indicate that cadmium is taken up by calcium and iron transporters, whereas chromium appears to enter the algae cells via iron and zinc carriers. It was shown that lead is not taken up into Micrasterias at all but exerts its adverse effects on cell growth by substituting cell wall bound calcium. The antioxidants salicylic acid, ascorbic acid and glutathione were not able to ameliorate any of the investigated metal effects on the green alga Micrasterias when added to the culture medium.

Intracellular chromium localization and cell physiological response in the unicellular alga Micrasterias.

Various contaminants like metals and heavy metals are constantly released into the environment by anthropogenic activities. The heavy metal chromium has a wide industrial use and exists in two stable oxidation states: trivalent and hexavalent. Chromium can cause harm to cell metabolism and development, when it is taken up by plants instead of necessary micronutrients such as for example iron. The uptake of Cr VI into plant cells has been reported to be an active process via carriers of essential anions, while the cation Cr III seems to be taken up inactively. Micrasterias denticulata, an unicellular green alga of the family Desmidiaceae is a well-studied cell biological model organism. Cr III and VI had inhibiting effects on its cell development, while cell division rates were only impaired by Cr VI. Transmission electron microscopy (TEM) revealed ultrastructural changes such as increased vacuolization, condensed cytoplasm and dark precipitations in the cell wall after 3 weeks of Cr VI treatment. Electron energy loss spectroscopy (EELS) and electron spectroscopic imaging (ESI) were applied to measure intracellular chromium distribution. Chromium was only detected after 3 weeks of 10 µM Cr VI treatment in electron dense precipitations found in bag-like structures along the inner side of the cell walls together with iron and elevated levels of oxygen, pointing toward an accumulation respectively extrusion of chromium in form of an iron-oxygen compound. Atomic emission spectroscopy (EMS) revealed that Micrasterias cells are able to accumulate considerable amounts of chromium and iron. During chromium treatment the Cr:Fe ratio shifted in favor of chromium, which implied that chromium may be taken up instead of iron. Significant and rapid increase of ROS production within the first 5 min of treatment confirms an active Cr VI uptake. SOD and CAT activity after Cr VI treatment did not show a response, while the glutathione pool determined by immuno-TEM decreased significantly in chromium treated cells, showing that glutathione is playing a major role in intracellular ROS and chromium detoxification.