International Journal of Molecular Sciences / Subcellular sequestration and impact of heavy metals on the ultrastructure and physiology of the multicellular freshwater alga Desmidium swartzii

Due to modern life with increasing traffic, industrial production and agricultural practices, high amounts of heavy metals enter ecosystems and pollute soil and water. As a result, metals can be accumulated in plants and particularly in algae inhabiting peat bogs of low pH and high air humidity. In the present study, we investigated the impact and intracellular targets of aluminum, copper, cadmium, chromium VI and zinc on the filamentous green alga Desmidium swartzii, which is an important biomass producer in acid peat bogs. By means of transmission electron microscopy (TEM) and electron energy loss spectroscopy (EELS) it is shown that all metals examined are taken up into Desmidium readily, where they are sequestered in cell walls and/or intracellular compartments. They cause effects on cell ultrastructure to different degrees and additionally disturb photosynthetic activity and biomass production. Our study shows a clear correlation between toxicity of a metal and the ability of the algae to compartmentalize it intracellularly. Cadmium and chromium, which are not compartmentalized, exert the most toxic effects. In addition, this study shows that the filamentous alga Desmidium reacts more sensitively to aluminum and zinc when compared to its unicellular relative Micrasterias, indicating a severe threat to the ecosystem.(VLID)156767

Gold nanoparticles (AuNPs) impair LPS-driven immune responses by promoting a tolerogenic-like dendritic cell phenotype with altered endosomal structures

Dendritic cells (DCs) shape immune responses by influencing T-cell activation. Thus, they are considered both an interesting model for studying nano-immune interactions and a promising target for nano-based biomedical applications. However, the accentuated ability of nanoparticles (NPs) to interact with biomolecules may have an impact on DC function that poses an unexpected risk of unbalanced immune reactions. Here, we investigated the potential effects of gold nanoparticles (AuNPs) on DC function and the consequences for effector and memory T-cell responses in the presence of the microbial inflammatory stimulus lipopolysaccharide (LPS). Overall, we found that, in the absence of LPS, none of the tested NPs induced a DC response. However, whereas 4-, 8-, and 11 nm AuNPs did not modulate LPS-dependent immune responses, 26 nm AuNPs shifted the phenotype of LPS-activated DCs toward a tolerogenic state, characterized by downregulation of CD86, IL-12 and IL-27, upregulation of ILT3, and induction of class E compartments. Moreover, this DC phenotype was less proficient in promoting Th1 activation and central memory T-cell proliferation. Taken together, these findings support the perception that AuNPs are safe under homeostatic conditions; however, particular care should be taken in patients experiencing a current infection or disorders of the immune system

Transcriptional analysis of cell growth and morphogenesis in the unicellular green alga Micrasterias (Streptophyta), with emphasis on the role of expansin

<p>Abstract</p> <p>Background</p> <p>Streptophyte green algae share several characteristics of cell growth and cell wall formation with their relatives, the embryophytic land plants. The multilobed cell wall of <it>Micrasterias denticulata </it>that rebuilds symmetrically after cell division and consists of pectin and cellulose, makes this unicellular streptophyte alga an interesting model system to study the molecular controls on cell shape and cell wall formation in green plants.</p> <p>Results</p> <p>Genome-wide transcript expression profiling of synchronously growing cells identified 107 genes of which the expression correlated with the growth phase. Four transcripts showed high similarity to expansins that had not been examined previously in green algae. Phylogenetic analysis suggests that these genes are most closely related to the plant EXPANSIN A family, although their domain organization is very divergent. A GFP-tagged version of the expansin-resembling protein MdEXP2 localized to the cell wall and in Golgi-derived vesicles. Overexpression phenotypes ranged from lobe elongation to loss of growth polarity and planarity. These results indicate that MdEXP2 can alter the cell wall structure and, thus, might have a function related to that of land plant expansins during cell morphogenesis.</p> <p>Conclusions</p> <p>Our study demonstrates the potential of <it>M. denticulata </it>as a unicellular model system, in which cell growth mechanisms have been discovered similar to those in land plants. Additionally, evidence is provided that the evolutionary origins of many cell wall components and regulatory genes in embryophytes precede the colonization of land.</p

The recombinant inhibitor of DNA binding Id2 forms multimeric structures via the helix-loop-helix domain and the nuclear export signal

The inhibitor of DNA binding and cell differentiation 2 (Id2) is a helix-loop-helix (HLH) protein that acts as negative dominant regulator of basic-HLH transcription factors during development and in cancer. The structural properties of Id2 have been investigated so far by using synthetic or recombinant fragments reproducing single domains (N-terminus, HLH, C-terminus): the HLH domain tends to dimerize into a four-helix bundle, whereas the flanking regions are flexible. In this work, the intact protein was expressed in E. coli, solubilized from inclusion bodies with urea, purified and dissolved in water at pH4. Under these conditions, Id2 was obtained with both cysteine residues disulfide-bonded to -mercaptoethanol that was present during the solubilization process. Moreover, it existed in a self-assembled state, in which the N-terminus remained highly flexible, while the HLH domain and, surprisingly, part of the C-terminus, which corresponds to the nuclear export signal (NES), both were involved in slowly tumbling, rigid structures. The protein oligomers also formed twisted fibrils that were several micrometers long and up to 80 nm thick. These results show that self-assembly decreases the backbone flexibility of those two protein regions (HLH and NES) that are important for interaction with basic-HLH transcription factors or for nucleocytoplasmic shuttling.(VLID)253307

Analyses and localization of pectin-like carbohydrates in cell wall and mucilage of the green alga Netrium digitus

The unicellular, simply shaped desmid Netrium digitus inhabiting acid bog ponds grows in two phases. Prior to division, the cell elongates at its central zone, whereas in a second phase, polar tip growth occurs. Electron microscopy demonstrates that Netrium is surrounded by a morphologically homogeneous cell wall, which lacks pores. Immunocytochemical and biochemical analyses give insight into physical wall properties and, thus, into adaptation to the extreme environment. The monoclonal antibodies JIM5 and JIM7 directed against pectic epitopes with different degrees of esterification label preferentially growing wall zones in Netrium. In contrast, 2F4 marks the cell wall only after experimental de-esterification. Electron energy loss spectroscopy reveals Ca-binding capacities of pectins and gives indirect evidence for the degree of their esterification. An antibody raised against Netrium mucilage is not only specific to mucilage but also recognizes wall components in transmission electron microscopy and dot blots. These results indicate a smooth transition between mucilage and the cell wall in Netrium

Chitosan functionalisation of gold nanoparticles encourages particle uptake and induces cytotoxicity and pro-inflammatory conditions in phagocytic cells, as well as enhancing particle interactions with serum components

Gold nanoparticles (AuNPs) are a popular choice for use in medical and biomedical research applications. With suitable functionalisation AuNPs can be applied in drug delivery systems, or can aid in disease diagnosis. One such functionalisation is with chitosan, which enables efficient interaction and permeation of cellular membranes, providing an effective adjuvant. As both AuNPs and chitosan have been shown to have low toxicity and high biocompatibility their proposed use in nanomedicine, either individually or combined, is expanding. However, further toxicological and immunological assessments of AuNP-chitosan conjugates are still needed. Therefore, we have evaluated how AuNP functionalisation with chitosan can affect uptake, cytotoxicity, and immunological responses within mononuclear cells, and influence the interaction of AuNPs with biomolecules within a complex biofluid. The AuNPs used were negatively charged through citrate-coating, or presented either low or high positive charge through chitosan-functionalisation. Uptake by THP-1 cells was assessed via transmission electron microscopy and electron energy loss spectroscopy, pro-inflammatory responses by ELISA and qRT-PCR, and cell death and viability via lactate dehydrogenase release and mitochondrial activity, respectively. Interactions of AuNPs with protein components of a frequently used in vitro cell culture medium supplement, foetal calf serum, were investigated using mass spectrometry. Although cells internalised all AuNPs, uptake rates and specific routes of intracellular trafficking were dependent upon chitosan-functionalisation. Accordingly, an enhanced immune response was found to be chitosan-functionalisation-dependent, in the form of CCL2, IL-1β, TNF-α and IL-6 secretion, and expression of IL - 1β and NLRP3 mRNA. A corresponding increase in cytotoxicity was found in response to chitosan-coated AuNPs. Furthermore, chitosan-functionalisation was shown to induce an increase in unique proteins associating with these highly charged AuNPs. It can be concluded that functionalisation of AuNPs with the perceived non-toxic biocompatible molecule chitosan at a high density can elicit functionalisation-dependent intracellular trafficking mechanisms and provoke strong pro-inflammatory conditions, and that a high affinity of these NP-conjugates for biomolecules may be implicit in these cellular responses. The online version of this article (doi:10.1186/s12951-015-0146-9) contains supplementary material, which is available to authorized users

Hyposaline conditions affect UV susceptibility in the Arctic kelp Alaria esculenta (Phaeophyceae) - results of laboratory experiments at Kongsfjorden, June/July 2014

The kelp Alaria esculenta represents a key species in high Arctic marine fjord ecosystems. However, the European Arctic is currently experiencing extensive environmental change. Glacial fjord systems, such as Kongsfjorden (Spitsbergen, Svalbard), are subjected to rising temperature, increased freshwater inflow from glaciers and melting snow and a changing ultraviolet (UV) radiation regime related to stratospheric ozone depletion. Thus, in addition to natural seasonality, sessile organisms require acclimation in order to adapt to an environment in transition. We examined the physiological and ultrastructural responses of A. esculenta to the combined exposure to hyposalinity and UV radiation. Photosynthetic quantum yield slightly decreased during a low-salinity treatment of 7 d. Exposure to UV radiation also lowered quantum yield, but specimens previously treated with hyposalinity were significantly less susceptible to UV than nontreated individuals. Concomitant with a loss of chlorophyll during the hyposaline treatment, phlorotannin and antioxidant contents were maintained, and samples treated with low salinities exhibited higher UV-screening characteristics as demonstrated by significantly higher absorption ratios at 300/680 nm. Ultrastructural analyses revealed a treatment-dependent swelling of cell walls and accumulations of phlorotannin-containing vesicles. Our findings point to a strategy by which kelps apply a fast and cost-efficient redistribution of phlorotannins rather than increased synthesis as a general stress response to different environmental drivers in contrast to stress-specific responses. The notion that acclimation to one stressor (low salinity) reflects increased tolerance towards a second stressor (UV radiation) supports the concept of 'cross-acclimation' as established for higher plants but not yet for seaweeds