Tracing the slow growth of anaerobic methane-oxidizing communities by 15N-labelling techniques

The anaerobic oxidation of methane (AOM) is an important methane sink in marine ecosystems mediated by still uncultured Archaea . We established an experimental system to grow AOM communities in different sediment samples. Approaches to show growth of the slow-growing anaerobic methanotrophs have been either via nucleic acids (quantitative PCR) or required long-term incubations. Previous long-term experiments with ¹³C-labelled methane led to an unspecific distribution of the ¹³C-label. Although quantitative PCR is a sensitive technique to detect small changes in community composition, it does not determine growth yield. Therefore, we tested an alternative method to detect a biomass increase of AOM microorganisms with ¹⁵N-labelled ammonium as N-source. After only 3 weeks, significant ¹⁵N-labelling became apparent in amino acids as major structural units of microbial proteins. This was especially evident in methane-containing incubations, showing the methane-dependent uptake of the ¹⁵N-labelled ammonium by microorganisms. Cell counts demonstrated a two- and fourfold increase at ambient or elevated methane concentrations. With denaturing gradient gel electrophoresis, over 6 months incubation no changes in community composition of sulphate-reducing bacteria and archaea were detected. These data indicate doubling times for AOM microorganisms between 2 and 3.4 months. In conclusion, the ¹⁵N-labelling approach proved to be a sensitive and fast way to show growth of extremely slow-growing microorganisms.     

Tunneling nanotubes,an emerging intercellular communication route in development

The development of multi-cellular organisms involves a comprehensive and tightly regulated cell-to-cell communication system to coordinate the activity and behavior of individual cells. Diverse signaling pathways ranging from receptor-mediated signal transduction to contact-dependent communication via gap junctions achieve these complex interactions. In this review, we will focus on a new type of intercellular connection, the tunneling nanotube (TNT), which has been observed in many cell types in vitro and recently also in developing embryos of different species in vivo. We will summarize the latest insights into their functional roles in cell-to-cell signaling with a particular focus on the TNT-dependent electrical coupling between developing embryonic cells. Finally, potential implications of these new findings in the light of developmental processes, particularly in cell migration, will be discussed.     

The role of myosin V in exocytosis and synaptic plasticity

In neuroscience, myosin V motor proteins have attracted attention since they are highly expressed in brain, and absence of myosin Va in man leads to a severe neurological disease called Griscelli syndrome. While in some cells myosin V is described to act as a vesicle transport motor, an additional role in exocytosis has emerged recently. In neurons, myosin V has been linked to exocytosis of secretory vesicles and recycling endosomes. Through these functions, it is implied in regulating important brain functions including the release of neuropeptides by exocytosis of large dense-core vesicles and the insertion of neurotransmitter receptors into post-synaptic membranes. This review focuses on the role of myosin V in (i) axonal transport and stimulated exocytosis of large dense-core vesicles to regulate the secretion of neuroactive substances, (ii) tethering of the endoplasmic reticulum at cerebellar synapses to permit long-term depression, (iii) recycling of α-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptors at hippocampal synapses during long-term potentiation, and (iv) recycling of nicotinic acetylcholine receptors at the neuromuscular junction. Myosin V is thus discussed as an important modulator of synaptic plasticity.     

Sulfidic acetate mineralization at 45°C by an aquifer microbial community: key players and effects of heat changes on activity and community structure

High-Temperature Aquifer Thermal Energy Storage (HT-ATES) is a sustainable approach for integrating thermal energy from various sources into complex energy systems. Temperatures ≥45°C, which are relevant in impact zones of HT-ATES systems, may dramatically influence the structure and activities of indigenous aquifer microbial communities. Here, we characterized an acetate-mineralizing, sulfate-reducing microbial community derived from an aquifer and adapted to 45°C. Acetate mineralization was strongly inhibited at temperatures ≤25°C and 60°C. Prolonged incubation at 12°C and 25°C resulted in acetate mineralization recovery after 40–80 days whereas acetate was not mineralized at 60°C within 100 days. Cultures pre-grown at 45°C and inhibited for 28 days by incubation at 12°C, 25°C, or 60°C recovered quickly after changing the temperature back to 45°C. Phylotypes affiliated to the order Spirochaetales and to endospore-forming sulfate reducers of the order Clostridiales were highly abundant in microcosms being active at 45°C highlighting their key role. In summary, prolonged incubation at 45°C resulted in active microbial communities mainly consisting of organisms adapted to temperatures between the typical temperature range of mesophiles and thermophiles and being resilient to temporary heat changes.     

miR-184 Regulates Pancreatic β-Cell Function According to Glucose Metabolism

In response to fasting or hyperglycemia, the pancreatic β-cell alters its output of secreted insulin; however, the pathways governing this adaptive response are not entirely established. Although the precise role of microRNAs (miRNAs) is also unclear, a recurring theme emphasizes their function in cellular stress responses. We recently showed that miR-184, an abundant miRNA in the β-cell, regulates compensatory proliferation and secretion during insulin resistance. Consistent with previous studies showing miR-184 suppresses insulin release, expression of this miRNA was increased in islets after fasting, demonstrating an active role in the β-cell as glucose levels lower and the insulin demand ceases. Additionally, miR-184 was negatively regulated upon the administration of a sucrose-rich diet in Drosophila, demonstrating strong conservation of this pathway through evolution. Furthermore, miR-184 and its target Argonaute2 remained inversely correlated as concentrations of extracellular glucose increased, underlining a functional relationship between this miRNA and its targets. Lastly, restoration of Argonaute2 in the presence of miR-184 rescued suppression of miR-375-targeted genes, suggesting these genes act in a coordinated manner during changes in the metabolic context. Together, these results highlight the adaptive role of miR-184 according to glucose metabolism and suggest the regulatory role of this miRNA in energy homeostasis is highly conserved.     

Effects of hydrogen and acetate on benzene mineralisation under sulphate-reducing conditions

Syntrophic mineralisation of benzene, as recently proposed for a sulphate-reducing enrichment culture, was tested in product inhibition experiments with acetate and hydrogen, both putative intermediates of anaerobic benzene fermentation. Using [(13)C(6)]-benzene enabled tracking the inhibition of benzene mineralisation sensitively by analysis of (13)CO(2). In noninhibited cultures, hydrogen was detected at partial pressures of 2.4 × 10(-6) ± 1.5 × 10(-6) atm. Acetate was detected at concentrations of 17 ± 2 μM. Spiking with 0.1 atm hydrogen produced a transient inhibitory effect on (13)CO(2) formation. In cultures spiked with higher amounts of hydrogen, benzene mineralisation did not restart after hydrogen consumption, possibly due to the toxic effects of the sulphide produced. An inhibitory effect was also observed when acetate was added to the cultures (0.3, 3.5 and 30 mM). Benzene mineralisation resumed after acetate was degraded to concentrations found in noninhibited cultures, indicating that acetate is another key intermediate in anaerobic benzene mineralisation. Although benzene mineralisation by a single sulphate reducer cannot be ruled out, our results strongly point to an involvement of syntrophic interactions in the process. Thermodynamic calculations revealed that, under in situ conditions, benzene fermentation to hydrogen and acetate yielded a free energy change of ΔG'=-83.1 ± 5.6 kJ mol(-1). Benzene mineralisation ceased when ΔG' values declined below -61.3 ± 5.3 kJ mol(-1) in the presence of acetate, indicating that ATP-consuming reactions are involved in the pathway.     

Activation of myosin V-based motility and F-actin-dependent network formation of endoplasmic reticulum during mitosis

Putative myogenic and endothelial (myo-endothelial) cell progenitors were identified in the interstitial spaces of murine skeletal muscle by immunohistochemistry and immunoelectron microscopy using CD34 antigen. Enzymatically isolated cells were characterized by fluorescence-activated cell sorting on the basis of cell surface antigen expression, and were sorted as a CD34+ and CD45- fraction. Cells in this fraction were approximately 94% positive for Sca-1, and mostly negative (<3% positive) for CD14, 31, 49, 144, c-kit, and FLK-1. The CD34+/45- cells formed colonies in clonal cell cultures and colony-forming units displayed the potential to differentiate into adipocytes, endothelial, and myogenic cells. The CD34+/45- cells fully differentiated into vascular endothelial cells and skeletal muscle fibers in vivo after transplantation. Immediately after sorting, CD34+/45- cells expressed only c-met mRNA, and did not express any other myogenic cell-related markers such as MyoD, myf-5, myf-6, myogenin, M-cadherin, Pax-3, and Pax-7. However, after 3 d of culture, these cells expressed mRNA for all myogenic markers. CD34+/45- cells were distinct from satellite cells, as they expressed Bcrp1/ABCG2 gene mRNA (Zhou et al., 2001). These findings suggest that myo-endothelial progenitors reside in the interstitial spaces of mammalian skeletal muscles, and that they can potentially contribute to postnatal skeletal muscle growth.     

Carbon isotope fractionation during <Emphasis Type="Italic">cis</Emphasis>–<Emphasis Type="Italic">trans</Emphasis> isomerization of unsaturated fatty acids in <Emphasis Type="Italic">Pseudomonas putida</Emphasis>

The molecular mechanism of the unique cis to trans isomerization of unsaturated fatty acids in the solvent-tolerant bacterium Pseudomonas putida S12 was studied. For this purpose, the carbon isotope fractionation of the cis–trans isomerase was estimated. In resting cell experiments, addition of 3-nitrotoluene for activation of the cis–trans isomerase resulted in the conversion of the cis-unsaturated fatty acids into the corresponding trans isomers. For the conversion of C16:1 cis to its corresponding trans isomer, a significant fractionation was measured. The intensity of this fractionation strongly depended on the rate of cis–trans isomerization and the added concentration of 3-nitrotoluene, respectively. The presence of a significant fractionation provides additional indication for a transition from the sp² carbon linkage of the cis-double bond to an intermediate sp³ within an enzyme–substrate complex. The sp² linkage is reconstituted after rotation to the trans configuration has occurred. As cytochrome c plays a major role in the catabolism of Cti polypeptide, these findings favour a mechanism for the enzyme in which electrophilic iron (Fe³⁺), provided by a heme domain, removes an electron of the cis double bond thereby transferring the sp² linkage into sp³.