Gene‐Centric Metagenome Analysis Reveals Gene Clusters for Carbon Monoxide Conversion and Validates Isolation of a Clostridial Acetogen for C2 Chemical Production

Syngas fermentation is largely dependent on acetogens that occur in various anaerobic environmental samples including soil, sediment, and feces. Here the authors report the metagenomic isolation of acetogens for C2 chemical production from syngas. Screening acetogens for C2 chemical production typically involves detecting the presence of the Wood‐Ljungdahl Pathway for carbon monoxide conversion. The authors collect samples from river‐bed sediments potentially having conditions suitable for carbon monoxide‐converting anaerobes, and enrich the samples under carbon monoxide selection pressure. Changes in the microbial community during the experimental procedure are investigated using both amplicon and shotgun metagenome sequencing. Combined next‐generation sequencing techniques enabl in situ tracking of the major acetogenic bacterial group and lead to the discovery of a 16 kb of gene cluster for WLP. The authors isolat an acetogenic clostridial strain from the enrichment culture (strain H21‐9). The functional activity of H21‐9 is confirmed by its high level of production of C2 chemicals from carbon monoxide (77.4 mM acetate and 2.5 mM of ethanol). This approach of incorporating experimental enrichment with metagenomic analysis can facilitate the discovery of novel strains from environmental habitats by tracking target strains during the screening process, combined with validation of their functional activity.     

The role of in vitro cultivation on symbiotic trait and function variation in a single species of arbuscular mycorrhizal fungus

In vitro propagation of AM fungi using transformed root cultures (TRC) is commonly used to obtain pure AM fungal propagules for use in research and industry. Early observations indicate that such an artificial environment can alter traits and function of AM fungi over time. We hypothesized that increased in vitro cultivation may promote ruderal strategies in fungi by enhancing propagule production and reducing mutualistic quality. To examine the effect of in vitro cultivation on the trait and function of AM fungi, we inoculated plants with 11 Rhizoglomus irregulare isolates which fell along a cultivation gradient spanning 80 generations. We harvested plants at 10, 20 and 30 d post inoculation to observe differences in fungal and plant traits post infection. In vitro cultivation led to increased spore production but reduced plant shoot phosphorus. Our results indicate that in vitro propagation may indirectly select for traits that affect symbiotic quality.     

Homeostatic Intrinsic Plasticity Is Functionally Altered in Fmr1 KO Cortical Neurons

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Modifying dipalmitoylphosphatidylcholine monolayers by n-hexadecanol and dipalmitoylglycerol

The monolayer structure of pure dipalmitoylphosphatidylcholine (DPPC) and equimolar mixtures of DPPC/n-hexadecanol (C(16)OH) and DPPC/dipalmitoylglycerol (DPG) are studied by the film balance technique and grazing incidence X-ray diffraction measurements. At 20 degrees C, the binary systems exhibit complete miscibility. In contrast to pure DPPC monolayers, a condensing effect is observed in the presence of both non-phospholipid additives; but the phase transition behavior differs. The tilt angle of the hydrocarbon chains in the DPPC/C(16)OH mixture is significantly smaller than in pure DPPC monolayers. The tilt of the chains is even further reduced in the mixed monolayer of DPPC/DPG. A comparison of the three systems reveals distinct structural features such as phase state, chain tilt, and molecular area over a wide range of surface pressures. Therefore, these monolayers provide a highly suitable model to investigate the influence of structural parameters on biological processes occurring at the membrane surface, e.g. enzymatic reactions and adsorption events.     

Heterogenecity of stromal cell precursers isolated from rat bone marrow

Bone marrow stroma contains mesenchymal stem cells (MSC) which are progenitor cells, at least for tissues arising from mesechyma. The study of MSC biology yields controversial data. Therefore further experiments are needed to characterize these cells. The aim of our research was to compare primary cultures and subcultures of stromal precursor cells isolated from rat bone marrow. Long-term cultures of these cells isolated from 5 animals have been obtained. Morphological, immunophenotypic, and functional (capacity to osteogenic differentiation) characteristics of the cells have been investigated. We show that the cell morphology in the cultures is highly heterogenic. Morphological cell types are described. Heterogeneity of stromal cells declines on late passages. Cell cultures isolated from different animals have the same immunophenotypic markers (CD90, CD44, CD54, CD106, CD45, CD11b) but different morphological characteristics and a different capacity to osteogenic differentiation during long-term cultivation. The data show that more specific markers and functional tests should be applied to identify MSC.     

Cellular and molecular control of dendritic growth and development of cerebellar Purkinje cells

Purkinje cells are the principal neurons of the cerebellar cortex and are characterized by a large and highly branched dendritic tree. For this reason, they have for a long time been an attractive model system to study the regulation of dendritic growth and differentiation. In this article, I will first review studies on different aspects of Purkinje cell dendritic development and then go on to present studies which have aimed at experimentally altering Purkinje cell dendritic development. Some of the cellular and molecular mechanisms which have been shown by these studies to be important determinants of Purkinje cell dendritic development will be discussed, in particular the role of the parallel fiber input, of hormones, and of neuronal growth factors. The organotypic slice culture method will be introduced as an important experimental tool to study Purkinje cell dendritic development under controlled conditions. Using cerebellar slice cultures, protein kinase C (PKC) has been identified as a major determinant of Purkinje cell dendritic development and the contribution of specific isoforms of PKC will be discussed. Finally, it will be shown that Purkinje cell dendritic development in slice cultures does not depend on the activation of glutamate receptors and appears to be independent of the presence of the neurotrophin BDNF. These studies indicate that the initial outgrowth of the Purkinje cell dendritic tree can occur in the absence of signals derived from afferent fibers, but is under control of PKC signaling.     

Astrocyte Mitochondria in In Vitro Models of Ischemia

There is growing evidence that preservation of mitochondrial respiratory function during cerebral ischemia-reperfusion predicts the ultimate extent of tissue injury. Because neurons are selectively vulnerable to ischemic injury, many studies have focused on neuronal mitochondrial dysfunction in ischemia. However, positron emission tomography (PET) studies in animals and humans suggest that non-neuronal cells such as astrocytes may also experience mitochondrial metabolic compromise that contributes to ischemic necrosis. Astrocytes carry out a number of functions that are critical to normal nervous system function, including uptake of neurotransmitters, regulation of pH and ion concentrations, and metabolic support of neurons. Mitochondria are important for many of these actions. We have used a cell culture model of stroke, oxygen-glucose deprivation (OGD), to study the response of astrocyte mitochondria to ischemia, and to evaluate how changes in astrocyte mitochondrial function might affect neuronal survival and recovery after ischemia.     

Purification,Cloning and Functional Expression of hydroxyphenylpyruvate Reductase Involved in Rosmarinic Acid Biosynthesis in Cell Cultures of Coleus Blumei

Hydroxyphenylpyruvate reductase (HPPR) is an enzyme involved in the biosynthesis of rosmarinic acid in Lamiaceae reducing hydroxyphenylpyruvates in dependence of NAD(P)H to the corresponding hydroxyphenyllactates. The HPPR protein was purified from suspension cells of Coleus blumei accumulating high levels of rosmarinic acid by ammonium sulfate precipitation, anion exchange chromatography, hydroxylapatite chromatography, chromatography on 2',5'-ADP-Sepharose 4B and SDS-polyacrylamide gel electrophoresis. The protein was tryptically digested and the peptides sequenced. Sequence information was used to isolate a full-length cDNA-clone for HPPR (EMBL accession number AJ507733) by RT-PCR, screening of a C. blumei cDNA-library and 5'-RACE-PCR. The open reading frame of the HPPR-cDNA consists of 939 nucleotides encoding a protein of 313 amino acid residues. The sequence showed that HPPR belongs to the family of D-isomer-specific 2-hydroxyacid dehydrogenases. The HPPR-cDNA was heterologously expressed in Escherichia coli and the protein was shown to catalyse the NAD(P)H-dependent reduction of 4-hydroxyphenylpyruvate to 4-hydroxyphenyllactate and 3,4-dihydroxyphenylpyruvate to 3,4-dihydroxyphenyllactate.     

Role of thioredoxin peroxidase in aging of stationary cultures of Saccharomyces cerevisiae

A soluble protein from Saccharomyces cerevisiae acts as a peroxidase but requires a NADPH-dependent thioredoxin system and was named thioredoxin peroxidase (TPx). The role of TPx in aging of stationary cultures of S. cerevisiae was investigated in a wild-type strain and a mutant yeast strain in which the tsa gene that encodes TPx was disrupted by homologous recombination. The occurrence of oxidative stress during aging of stationary cultures of the yeast has been proposed. Comparison of 5-day-old (young) stationary cultures of S. cerevisiae and of cultures aged for 3 months (old) revealed decreased viability, increased generation of reactive oxygen species, modulation of cellular redox status, and increased cellular oxidative damage reflected by increased protein carbonyl content and lipid peroxidation. The magnitude of this stress was augmented in yeast mutant lacking TPx. These results suggest that TPx may play a direct role in cellular defense against aging of stationary cultures presumably, functioning as an antioxidant enzyme.