SERF protein is a direct modifier of amyloid fiber assembly

The inherent cytotoxicity of aberrantly folded protein aggregates contributes substantially to the pathogenesis of amyloid diseases. It was recently shown that a class of evolutionary conserved proteins, called MOAG-4/SERF, profoundly alter amyloid toxicity via an autonomous but yet unexplained mode. We show that the biological function of human SERF1a originates from its atypical ability to specifically distinguish between amyloid and nonamyloid aggregation. This inherently unstructured protein directly affected the aggregation kinetics of a broad range of amyloidogenic proteins in vitro, while being inactive against nonamyloid aggregation. A representative biophysical analysis of the SERF1a:α-synuclein (aSyn) complex revealed that the amyloid-promoting activity resulted from an early and transient interaction, which was sufficient to provoke a massive increase of soluble aSyn amyloid nucleation templates. Therefore, the autonomous amyloid-modifying activity of SERF1a observed in living organisms relies on a direct and dedicated manipulation of the early stages in the amyloid aggregation pathway.     

Methane-Carbon Flow into the Benthic Food Web at Cold Seeps – A Case Study from the Costa Rica Subduction Zone

Cold seep ecosystems can support enormous biomasses of free-living and symbiotic chemoautotrophic organisms that get their energy from the oxidation of methane or sulfide. Most of this biomass derives from animals that are associated with bacterial symbionts, which are able to metabolize the chemical resources provided by the seeping fluids. Often these systems also harbor dense accumulations of non-symbiotic megafauna, which can be relevant in exporting chemosynthetically fixed carbon from seeps to the surrounding deep sea. Here we investigated the carbon sources of lithodid crabs (Paralomis sp.) feeding on thiotrophic bacterial mats at an active mud volcano at the Costa Rica subduction zone. To evaluate the dietary carbon source of the crabs, we compared the microbial community in stomach contents with surface sediments covered by microbial mats. The stomach content analyses revealed a dominance of epsilonproteobacterial 16S rRNA gene sequences related to the free-living and epibiotic sulfur oxidiser Sulfurovum sp. We also found Sulfurovum sp. as well as members of the genera Arcobacter and Sulfurimonas in mat-covered surface sediments where Epsilonproteobacteria were highly abundant constituting 10% of total cells. Furthermore, we detected substantial amounts of bacterial fatty acids such as i-C15∶0 and C17∶1ω6c with stable carbon isotope compositions as low as −53‰ in the stomach and muscle tissue. These results indicate that the white microbial mats at Mound 12 are comprised of Epsilonproteobacteria and that microbial mat-derived carbon provides an important contribution to the crab''s nutrition. In addition, our lipid analyses also suggest that the crabs feed on other ¹³C-depleted organic matter sources, possibly symbiotic megafauna as well as on photosynthetic carbon sources such as sedimentary detritus.     

Enterohemorrhagic Escherichia coli Hemolysin Employs Outer Membrane Vesicles to Target Mitochondria and Cause Endothelial and Epithelial Apoptosis

Enterohemorrhagic Escherichia coli (EHEC) strains cause diarrhea and hemolytic uremic syndrome resulting from toxin-mediated microvascular endothelial injury. EHEC hemolysin (EHEC-Hly), a member of the RTX (repeats-in-toxin) family, is an EHEC virulence factor of increasingly recognized importance. The toxin exists as free EHEC-Hly and as EHEC-Hly associated with outer membrane vesicles (OMVs) released by EHEC during growth. Whereas the free toxin is lytic towards human endothelium, the biological effects of the OMV-associated EHEC-Hly on microvascular endothelial and intestinal epithelial cells, which are the major targets during EHEC infection, are unknown. Using microscopic, biochemical, flow cytometry and functional analyses of human brain microvascular endothelial cells (HBMEC) and Caco-2 cells we demonstrate that OMV-associated EHEC-Hly does not lyse the target cells but triggers their apoptosis. The OMV-associated toxin is internalized by HBMEC and Caco-2 cells via dynamin-dependent endocytosis of OMVs and trafficked with OMVs into endo-lysosomal compartments. Upon endosome acidification and subsequent pH drop, EHEC-Hly is separated from OMVs, escapes from the lysosomes, most probably via its pore-forming activity, and targets mitochondria. This results in decrease of the mitochondrial transmembrane potential and translocation of cytochrome c to the cytosol, indicating EHEC-Hly-mediated permeabilization of the mitochondrial membranes. Subsequent activation of caspase-9 and caspase-3 leads to apoptotic cell death as evidenced by DNA fragmentation and chromatin condensation in the intoxicated cells. The ability of OMV-associated EHEC-Hly to trigger the mitochondrial apoptotic pathway in human microvascular endothelial and intestinal epithelial cells indicates a novel mechanism of EHEC-Hly involvement in the pathogenesis of EHEC diseases. The OMV-mediated intracellular delivery represents a newly recognized mechanism for a bacterial toxin to enter host cells in order to target mitochondria.     

The Bacterial SMC Complex Displays Two Distinct Modes of Interaction with the Chromosome

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A Crystallin Fold in the Interleukin-4-inducing Principle of Schistosoma mansoni Eggs (IPSE/α-1) Mediates IgE Binding for Antigen-independent Basophil Activation

The IL-4-inducing principle from Schistosoma mansoni eggs (IPSE/α-1), the major secretory product of eggs from the parasitic worm S. mansoni, efficiently triggers basophils to release the immunomodulatory key cytokine interleukin-4. Activation by IPSE/α-1 requires the presence of IgE on the basophils, but the detailed molecular mechanism underlying activation is unknown. NMR and crystallographic analysis of IPSEΔNLS, a monomeric IPSE/α-1 mutant, revealed that IPSE/α-1 is a new member of the βγ-crystallin superfamily. We demonstrate that this molecule is a general immunoglobulin-binding factor with highest affinity for IgE. NMR binding studies of IPSEΔNLS with the 180-kDa molecule IgE identified a large positively charged binding surface that includes a flexible loop, which is unique to the IPSE/α-1 crystallin fold. Mutational analysis of amino acids in the binding interface showed that residues contributing to IgE binding are important for IgE-dependent activation of basophils. As IPSE/α-1 is unable to cross-link IgE, we propose that this molecule, by taking advantage of its unique IgE-binding crystallin fold, activates basophils by a novel, cross-linking-independent mechanism.     

Ultrastructure of four types of striated muscle fibers in the atlantic hagfish (Myxine glutinosa,L.)

Summary Four types of striated muscle fibers with distinctive ultrastructure were defined in the Atlantic hagfish (Myxine glutinosa, L.): white, intermediate, and red fibers of m. parietalis, and red fibers of m. craniovelaris.White fibers are thick, contain very few mitochondria and fat vacuoles, and possess distinct and separate myofibrils with thin Z-disks and distinct M-lines. Intermediate fibers are thinner, possess largely similar myofibrils that often are even better separated due to a higher content of fat vacuoles and especially mitochondria and glycogen granules. Red fibers of m. parietalis contain large amounts of mitochondria, fat vacuoles, and glycogen granules. Their myofibrils possess M-lines, and although branching more, the myofibrils of red fibers conform with a Fibrillenstruktur pattern like those of white and intermediate fibers. Red fibers of m. craniovelaris are very thin, possess many smaller fat vacuoles, and large amounts of mitochondria and glycogen granules. The myofibrils are significantly thinner than in m. parietalis fibers, run as quite independent well separated units, possess thicker Z-disks, and lack M-lines. Large amounts of myosatellite cells are associated with these red fibers.Triads are located near A/I-junctions in all four fiber types and occur irregularly, the density of triads being different in the various fiber types.We are indebted to Dr. Finn Walvig, Biological Station, University of Oslo, Drøbak, for supply of hagfishes, and we also wish to thank Dr. Jan K. S. Jansen, Institute of Physiology, University of Oslo, for valuable suggestions during this study.     

Dense-core vesicles in motor nerve terminals

Summary Motor nerve terminals on white and intermediate muscle fibers of the Atlantic hagfish (Myxine glutinosa, L.) contain translucent synpatic vesicles and about 1–2% dense-core vesicles. Terminals on red muscle fibers contain up to 40% dense-core vesicles with diameter 800–1100 Å. Examinations for formaldehyde-induced fluorescence indicate yellow fluorescence (5-HT ?) apparently corresponding with terminal axons on red muscle fibers in craniovelar muscles. Possibly red muscle fibers of Myxine receive monoaminergic innervation.The author is indebted to Dr. Finn Walvig, Biological station, University of Oslo, Drøbak, for supply of hagfishes.     

The pyridine nucleotide and non-pyridine nucleotide dependence of L-glutamate dehydrogenase in the histochemical system

Summary Under histochemical conditions (fresh frozen sections from liver, kidney and cerebellum of the rat) it was shown that the oxidation of L-glutamic acid was carried out by the NAD-dependent L-glutamate dehydrogenase (E.C. 1.4.1.2) and/or the NAD- or NADP-dependent L-glutamate dehydrogenase (E.C. 1.4.1.3) as well as by an enzyme system which is not dependent on externally added NAD, NADP, FAD, FMN or CoQ₁₀ for activity.This non-pyridine dependent activity was related to the L-glutamate dehydrogenases proper, owing to the following: a) the localization of activity noticed corresponds to that obtained with the NAD- or NADP-containing media, b) the incubation time needed for initial formation of red and blue formazans is similar to that with coenzyme-containing media, c) pre-extraction experiments reveal similarity in enzyme diffusion rates, d) the named activity is influenced by the same agents and to the same extent as the activity obtained by the inclusion of NAD or NADP (e.g. dissociation of the dehydrogenase molecule into subunits due to urea, inhibition of activity due to N-ethyl maleimide and 1.10-phenanthroline, activation due to the allosteric effect of ADP and to high substrate concentration, allosteric inhibition caused by GTP and inhibition caused by -ketoglutaric acid, no inhibitory effect of KCN), and e) the named activity was not affected by added PMS (excluding activity due to L-aminoacid oxidase).In the in situ localization of enzyme activity it was found that L-glutamate dehydrogenases E.C. 1.4.1.2 and E.C. 1.4.1.3 co-exist in the cells of kidney and cerebellum, while the L-glutamate dehydrogenase E.C. 1.4.1.3 only was present in liver cells.Finally, it was stated that incubation time should be kept as short as possible in order to avoid Nothing dehydrogenase reaction as well as inhibition due to accumulation of -ketoglutaric acid. Only gel incubation media should be applied.Recipient of a research grant from the Danish Ministry of Education     

Endosymbioses between bacteria and deep-sea siboglinid tubeworms from an Arctic Cold Seep (Haakon Mosby Mud Volcano, Barents Sea)

Siboglinid tubeworms do not have a mouth or gut and live in obligate associations with bacterial endosymbionts. Little is currently known about the phylogeny of frenulate and moniliferan siboglinids and their symbionts. In this study, we investigated the symbioses of two co-occurring siboglinid species from a methane emitting mud volcano in the Arctic Ocean (Haakon Mosby Mud Volcano, HMMV): Oligobrachia haakonmosbiensis (Frenulata) and Sclerolinum contortum (Monilifera). Comparative sequence analysis of the host-specific 18S and the symbiont-specific 16S rRNA genes of S. contortum showed that the close phylogenetic relationship of this host to vestimentiferan siboglinids was mirrored in the close relationship of its symbionts to the sulfur-oxidizing gammaproteobacterial symbionts of vestimentiferans. A similar congruence between host and symbiont phylogeny was observed in O. haakonmosbiensis: both this host and its symbionts were most closely related to the frenulate siboglinid O. mashikoi and its gammaproteobacterial symbiont. The symbiont sequences from O. haakonmosbiensis and O. mashikoi formed a clade unaffiliated with known methane- or sulfur-oxidizing bacteria. Fluorescence in situ hybridization indicated that the dominant bacterial phylotypes originated from endosymbionts residing inside the host trophosome. In both S. contortum and O. haakonmosbiensis, characteristic genes for autotrophy (cbbLM) and sulfur oxidation (aprA) were present, while genes diagnostic for methanotrophy were not detected. The molecular data suggest that both HMMV tubeworm species harbour chemoautotrophic sulfur-oxidizing symbionts. In S. contortum, average stable carbon isotope values of fatty acids and cholesterol of -43 per thousand were highly negative for a sulfur oxidizing symbiosis, but can be explained by a (13)C-depleted CO(2) source at HMMV. In O. haakonmosbiensis, stable carbon isotope values of fatty acids and cholesterol of -70 per thousand are difficult to reconcile with our current knowledge of isotope signatures for chemoautotrophic processes.     

Lipid body formation plays a central role in cell fate determination during developmental differentiation of Myxococcus xanthus

Cell differentiation is widespread during the development of multicellular organisms, but rarely observed in prokaryotes. One example of prokaryotic differentiation is the Gram-negative bacterium Myxococcus xanthus . In response to starvation, this gliding bacterium initiates a complex developmental programme that results in the formation of spore-filled fruiting bodies. How the cells metabolically support the necessary complex cellular differentiation from rod-shaped vegetative cells into spherical spores is unknown. Here, we present evidence that intracellular lipid bodies provide the necessary metabolic fuel for the development of spores. Formed at the onset of starvation, these lipid bodies gradually disappear until they are completely used up by the time the cells have become mature spores. Moreover, it appears that lipid body formation in M. xanthus is an important initial step indicating cell fate during differentiation. Upon starvation, two subpopulations of cells occur: cells that form lipid bodies invariably develop into spores, while cells that do not form lipid bodies end up becoming peripheral rods, which are cells that lack signs of morphological differentiation and stay in a vegetative-like state. These data indicate that lipid bodies not only fuel cellular differentiation but that their formation represents the first known morphological sign indicating cell fate during differentiation.