CpG and LPS can interfere negatively with prion clearance in macrophage and microglial cells

Cells of the innate immune system play important roles in the progression of prion disease after peripheral infection. It has been found in vivo and in vitro that the expression of the cellular prion protein (PrP(c)) is up-regulated on stimulation of immune cells, also indicating the functional importance of PrP(c) in the immune system. The aim of our study was to investigate the impact of cytosine-phosphate-guanosine- and lipopolysaccharide-induced PrP(c) up-regulation on the uptake and processing of the pathological prion protein (PrP(Sc)) in phagocytic innate immune cells. For this purpose, we challenged the macrophage cell line J774, the microglial cell line BV-2 and primary bone marrow-derived macrophages in a resting or stimulated state with various prion strains, and monitored the uptake and clearance of PrP(Sc). Interestingly, stimulation led either to a transient increase in the level of PrP(Sc) relative to unstimulated cells or to a decelerated degradation of PrP(Sc). These features were dependent on cell type and prion strain. Our data indicate that the stimulation of innate immune cells may be able to support transient prion propagation, possibly explained by an increased PrP(c) cell surface expression in stimulated cells. We suggest that stimulation of innate immune cells can lead to an imbalance between the propagation and degradation of PrP(Sc).     

Certification to ensure sustainable production of biofuels

Sustainability risks increase with rising biofuel production. Environmental, social and economic sustainability issues in agricultural production and conversion processes have become a major concern. External effects of biofuel production are not covered by a market mechanism. Therefore, an instrument to address the most pressing sustainability issues of biofuel production is required. The article provides the outline of an implementable certification concept that was elaborated in a multi-stakeholder approach with companies and organizations from Europe, the Americas, and Asia. The approach chosen for certification is beneficial for stakeholders, is market based and does not hamper international trade. The proposed certification system is focused on the most pressing sustainability issues, such as conversion of high carbon density and high nature value land. “Major must” and “minor must” criteria have been developed to assess the sustainability. The certification itself should occur through a compact and cost-effective system. A meta system approach will allow the use of already existing standards. Two separate certificates, one for sustainability of biomass production and one for greenhouse gas emissions, are proposed. The certificates are decoupled in a book and claim system from the biomass or biofuels traded and will be traded on a market place. The general concept of this certification system for biomass and biofuels has been discussed intensively with industry and trade companies and with public sector organizations and nongovernmental organizations. The practical feasibility should be tested in a pilot phase with co-operation of the stakeholders who are already involved.     

The proteasome maturation protein POMP facilitates major steps of 20S proteasome formation at the endoplasmic reticulum

The quality control of proteins mediated by the plasticity of the proteasome system is regulated by the timely and flexible formation of this multisubunit proteolytic enzyme complex. Adaptable biogenesis of the 20S proteasome core complex is therefore of vital importance for adjusting to changing proteolytic requirements. However, the molecular mechanism and the cellular sites of mammalian proteasome formation are still unresolved. By using precursor complex-specific antibodies, we now show that the main steps in 20S core complex formation take place at the endoplasmic reticulum (ER). Thereby, the proteasome maturation protein (POMP)--an essential factor of mammalian proteasome biogenesis--interacts with ER membranes, binds to alpha1-7 rings, recruits beta-subunits stepwise and mediates the association of mammalian precursor complexes with the ER. Thus, POMP facilitates the main steps in 20S core complex formation at the ER to coordinate the assembly process and to provide cells with freshly formed proteasomes at their site of function.     

Proteinases in the joint: clinical relevance of proteinases in joint destruction

Proteinases are involved in essential steps in cartilage and bone homeostasis. Consequently, efforts have been made to establish their potential role in the pathology of rheumatic conditions such as rheumatoid arthritis, osteoarthritis and spondyloarthritis. Matrix metalloproteinases (MMPs) are sensitive markers of disease severity and response to treatment, and therefore they have potential in the assessment of rheumatic diseases. Despite disappointing early results with synthetic inhibitors of MMPs, there is still much scope for developing effective and safe MMPs inhibitors, and consequently to deliver new options to inhibit joint destruction.     

The role of membrane lipids in the induction of macrophage apoptosis by microparticles

Microparticles are membrane-derived vesicles that are released from cells during activation or cell death. These particles can serve as mediators of intercellular cross-talk and induce a variety of cellular responses. Previous studies have shown that macrophages undergo apoptosis after phagocytosing microparticles. Here, we have addressed the hypothesis that microparticles trigger this process via lipid pathways. In these experiments, microparticles induced apoptosis in primary macrophage cells or cell lines (RAW 264.7 or U937) with up to a 5-fold increase. Preincubation of macrophages with phosphatidylinositol-3,5-bisphosphate (PtdIns(3,5)BP) reduced the microparticle-induced apoptosis in a dose-dependent manner. PtdIns(3,5)BP is a specific inhibitor of the acid sphingomyelinase and thus can block the generation of pro-apoptotic ceramides. Similarly, the pre-incubation of macrophages with PtdIns(3,5)BP prevented microparticle-induced upregulation of caspase 8, which is a major target molecule of ceramide action in the apoptosis pathway. PtdIns(3,5)BP, however, had no effect on the spontaneous rate of apoptosis. To evaluate further signaling pathways induced by microparticles, the extracellular signal regulated kinase (ERK-) 1 was investigated. This kinase plays a role in activating phospholipases A2 which cleaves membrane phospholipids into arachidonic acid; microparticles have been suggested to be a preferred substrate for phospholipases A2. As shown in our experiments, microparticles strongly increased the amount of phosphorylated ERK1/2 in RAW 264.7 macrophages in a time-dependent manner, peaking 15 min after co-incubation. Addition of PD98059, a specific inhibitor of ERK1, prevented the increase in apoptosis of RAW 264.7 macrophages. Together, these data suggest that microparticles perturb lipid homeostasis of macrophages and thereby induce apoptosis. These results emphasize the importance of biolipids in the cellular cross-talk of immune cells. Based on the fact that in clinical situations with excessive cell death such as malignancies, autoimmune diseases and following chemotherapies high levels of circulating microparticles might modulate phagocytosing cells, a suppression of the immune response might occur due to loss of macrophages.     

Differential thermal adaptation of clonal strains of a protist morphospecies originating from different climatic zones

Eco-physiological variation and local adaptation are key issues in microbial ecology. Here, we investigated the thermal adaptation of 19 strains of the same Spumella morphospecies (Chrysophyceae, Heterokonta). In order to test for local adaptation and the existence of specific ecotypes we analysed growth rates of these strains, which originated from different climate regions. We applied temperature-adaptation as an eco-physiological marker and analysed growth rates of the different Spumella strains at temperatures between 0 degrees C and 35 degrees C. The temperatures allowing for maximal growth of strains from temperate and warm climatic zones ranged between 19.9 degrees C and 33.4 degrees C. Phylogenetically, most of these 'warm'-adapted strains fall into two different previously defined 18S rDNA Spumella clusters, one of them consisting of mostly soil organisms and the other one being a freshwater cluster. As a rule, the 'warm'-adapted strains of the soil cluster grew slower than the 'warm'-adapted isolates within the freshwater cluster. This difference most probably reflect different strategies, i.e. the formation of cysts at the expense of lower growth rates in soil organisms. In contrast, as expected, all isolates from Antarctica were cold-adapted and grew already around melting point of freshwater. Surprisingly, optimum temperature for these strains was between 11.8 degrees C and 17.7 degrees C and maximum temperature tolerated was between 14.6 degrees C and 23.5 degrees C. Our data indicate that despite the relatively high optimal temperature of most Antarctic strains, they may have a relative advantage below 5-10 degrees C only. Based on the thermal adaptation of the flagellate strains the Antarctic strains were clearly separated from the other investigated strains. This may indicate a limited dispersal of flagellates to and from Antarctica. Even if the latter assumption needs support from more data, we argue that the high levels of eco-physiological and molecular microdiversity indicate that the current species concepts do not sufficiently reflect protist eco-physiological differentiation.     

Coping with cyclic oxygen availability: evolutionary aspects

Both the gradual rise in atmospheric oxygen over the ProterozoicEon as well as episodic fluctuations in oxygen over severalmillion-year time spans during the Phanerozoic Era, have arguablyexerted strong selective forces on cellular and organismic respiratoryspecialization and evolution. The rise in atmospheric oxygen,some 2 billion years after the origin of life, dramaticallyaltered cell biology and set the stage for the appearance ofmulticelluar life forms in the Vendian (Ediacaran) Period ofthe Neoproterozoic Era. Over much of the Paleozoic, the levelof oxygen in the atmosphere was near the present atmosphericlevel (21%). In the Late Paleozoic, however, there were extendedtimes during which the level of atmospheric oxygen was eithermarkedly lower or markedly higher than 21%. That these Paleozoicshifts in atmospheric oxygen affected the biota is suggestedby the correlations between: (1) Reduced oxygen and the occurrencesof extinctions, a lowered biodiversity and shifts in phyleticsuccession, and (2) During hyperoxia, the corresponding occurrenceof phenomena such as arthropod gigantism, the origin of insectflight, and the evolution of vertebrate terrestriality. Basicsimilarities in features of adaptation to hyopoxia, manifestin living organisms at levels ranging from genetic and cellularto physiological and behavioral, suggest the common and earlyorigin of a suite of adaptive mechanisms responsive to fluctuationsin ambient oxygen. Comparative integrative approaches addressingthe molecular bases of phenotypic adjustments to cyclic oxygenfluctuation provide broad insight into the incremental stepsleading to the early evolution of homeostatic respiratory mechanismsand to the specialization of organismic respiratory function.     

Expression of a ULK1/2 binding-deficient ATG13 variant can partially restore autophagic activity in ATG13-deficient cells

Autophagy describes an intracellular process responsible for the lysosome-dependent degradation of cytosolic components. The ULK1/2 complex comprising the kinase ULK1/2 and the accessory proteins ATG13, RB1CC1, and ATG101 has been identified as a central player in the autophagy network, and it represents the main entry point for autophagy-regulating kinases such as MTOR and AMPK. It is generally accepted that the ULK1 complex is constitutively assembled independent of nutrient supply. Here we report the characterization of the ATG13 region required for the binding of ULK1/2. This binding site is established by an extremely short peptide motif at the C terminus of ATG13. This motif is mandatory for the recruitment of ULK1 into the autophagy-initiating high-molecular mass complex. Expression of a ULK1/2 binding-deficient ATG13 variant in ATG13-deficient cells resulted in diminished but not completely abolished autophagic activity. Collectively, we propose that autophagy can be executed by mechanisms that are dependent or independent of the ULK1/2-ATG13 interaction.