Seipin regulates excitatory synaptic transmission in cortical neurons

Heterozygosity for missense mutations in Seipin, namely N88S and S90L, leads to a broad spectrum of motor neuropathy, while a number of loss‐of‐function mutations in Seipin are associated with the Berardinelli–Seip congenital generalized lipodystrophy type 2 (CGL2, BSCL2), a condition that is characterized by severe lipoatrophy, insulin resistance, and intellectual impairment. The mechanisms by which Seipin mutations lead to motor neuropathy, lipodystrophy, and insulin resistance, and the role Seipin plays in central nervous system (CNS) remain unknown. The goal of this study is to understand the functions of Seipin in the CNS using a loss‐of‐function approach, i.e. by knockdown (KD) of Seipin gene expression. Excitatory post‐synaptic currents (EPSCs) were impaired in Seipin‐KD neurons, while the inhibitory post‐synaptic currents (IPSCs) remained unaffected. Expression of a shRNA‐resistant human Seipin rescued the impairment of EPSC produced by Seipin KD. Furthermore, α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA)‐induced whole‐cell currents were significantly reduced in Seipin KD neurons, which could be rescued by expression of a shRNA‐resistant human Seipin. Fluorescent imaging and biochemical studies revealed reduced level of surface AMPA receptors, while no obvious ultrastructural changes in the pre‐synapse were found. These data suggest that Seipin regulates excitatory synaptic function through a post‐synaptic mechanism.     

Bacterial microbiome of Coptotermes curvignathus (Isoptera: Rhinotermitidae) reflects the coevolution of species and dietary pattern

Coptotermes curvignathus Holmgren is capable of feeding on living trees. This ability is attributed to their effective digestive system that is furnished by the termite's own cellulolytic enzymes and cooperative enzymes produced by their gut microbes. In this study, the identity of an array of diverse microbes residing in the gut of C. curvignathus was revealed by sequencing the near‐full‐length 16S rRNA genes. A total of 154 bacterial phylotypes were found. The Bacteroidetes was the most abundant phylum and accounted for about 65% of the gut microbial profile. This is followed by Firmicutes, Actinobacteria, Spirochetes, Proteobacteria, TM7, Deferribacteres, Planctomycetes, Verrucomicrobia, and Termite Group 1. Based on the phylogenetic study, this symbiosis can be a result of long coevolution of gut enterotypes with the phylogenic distribution, strong selection pressure in the gut, and other speculative pressures that determine bacterial biome to follow. The phylogenetic distribution of cloned rRNA genes in the bacterial domain that was considerably different from other termite reflects the strong selection pressures in the gut where a proportional composition of gut microbiome of C. curvignathus has established. The selection pressures could be linked to the unique diet preference of C. curvignathus that profoundly feeds on living trees. The delicate gut microbiome composition may provide available nutrients to the host as well as potential protection against opportunistic pathogen.     

Long-term operation of double chambered microbial fuel cell for bio-electro denitrification

The main aim of this study is to investigate the performance of organic oxidation and denitrification of the system under long-term operation. The MFC reactor was operated in continuous mode for 180 days. Nitrate was successfully demonstrated as terminal electron acceptor, where nitrate was reduced at the cathode using electron provided by acetate oxidation at the anode. The removal efficiencies of chemical oxygen demand (COD) and nitrate were higher in the closed circuit system than in open circuit system. Both COD and nitrate reduction improved with the increase of organic loading and subsequently contributed to higher power output. The maximum nitrate removal efficiency was 88 ± 4 % (influent of 141 ± 14 mg/L). The internal resistant was 50 Ω, which was found to be low for a double chambered MFC. The maximum power density was 669 mW/m³ with current density of 3487 mA/m³.     

PALMA: mRNA to genome alignments using large margin algorithms

MOTIVATION: Despite many years of research on how to properly align sequences in the presence of sequencing errors, alternative splicing and micro-exons, the correct alignment of mRNA sequences to genomic DNA is still a challenging task. RESULTS: We present a novel approach based on large margin learning that combines accurate splice site predictions with common sequence alignment techniques. By solving a convex optimization problem, our algorithm-called PALMA-tunes the parameters of the model such that true alignments score higher than other alignments. We study the accuracy of alignments of mRNAs containing artificially generated micro-exons to genomic DNA. In a carefully designed experiment, we show that our algorithm accurately identifies the intron boundaries as well as boundaries of the optimal local alignment. It outperforms all other methods: for 5702 artificially shortened EST sequences from Caenorhabditis elegans and human, it correctly identifies the intron boundaries in all except two cases. The best other method is a recently proposed method called exalin which misaligns 37 of the sequences. Our method also demonstrates robustness to mutations, insertions and deletions, retaining accuracy even at high noise levels. AVAILABILITY: Datasets for training, evaluation and testing, additional results and a stand-alone alignment tool implemented in C++ and python are available at http://www.fml.mpg.de/raetsch/projects/palma     

Progesterone reduces erectile dysfunction in sleep-deprived spontaneously hypertensive rats

Background

The rate-limiting step in prostaglandin (PG) biosynthesis is catalyzed by phospholipase A2 (PLA2) enzymes which hydrolyze arachidonic acid from membrane phospholipids. Despite their importance in uterine PG production, little is known concerning the specific PLA2 enzymes that regulate arachidonic acid liberation in the uterine endometrium. The objectives of this study were to evaluate the expression and activities of calcium-independent Group VI and Group IVC PLA2 (PLA2G6 and PLA2G4C) and calcium-dependent Group IVA PLA2 (PLA2G4A) enzymes in the regulation of bovine uterine endometrial epithelial cell PG production.

Methods

Bovine endometrial epithelial cells in culture were treated with oxytocin, interferon-tau and the PLA2G6 inhibitor bromoenol lactone, alone and in combination. Concentrations of PGF2alpha and PGE2 released into the medium were analyzed. Western blot analysis was performed on cellular protein to determine the effects of treatments on expression of PLA2G4A, PLA2G6 and PLA2G4C. Group-specific PLA2 activity assays were performed on cell lysates following treatment with oxytocin, interferon-tau or vehicle (control), alone and in combination. To further evaluate the role of specific PLA2 enzymes in uterine cell PG biosynthesis, cells were transfected with cDNAs encoding human PLA2G6 and PLA24C, treated as described above and PG assays performed.

Results

Constitutive cell production of PGF2alpha was about two-fold higher than PGE2. Oxytocin stimulated production of both PGs but the increase of PGF2alpha was significantly greater. Interferon-tau diminished oxytocin stimulation of both PGs. The PLA2G6 inhibitor, bromoenol lactone, abolished oxytocin-stimulated production of PGF2alpha. Treatments had little effect on PLA2G4A protein expression. In contrast, oxytocin enhanced expression of PLA2G6 and this effect was diminished in the presence of interferon-tau. Expression of PLA2G4C was barely detectable in control and oxytocin treated cells but it was enhanced in cells treated with interferon-tau. Oxytocin stimulated PLA2 activity in assays designed to evaluate PLA2G6 activity and interferon-tau inhibited this response. In assays designed to measure PLA2G4C activity, only interferon-tau was stimulatory. Cells overexpressing PLA2G6 produced similar quantities of the two PGs and these values were significantly higher than PG production by non-transfected cells. Oxytocin stimulated production of both PGs and this response was inhibited by interferon-tau. Bromoenol lactone inhibited oxtocin stimulation of PGF2alpha production but stimulated PGE2 production, both in the absence and presence of oxytocin. Cells over-expressing PLA2G4C produced more PGE2 than PGF2alpha and interferon-tau stimulated PGE2 production.

Conclusion

Results from these studies indicate that oxytocin stimulation of uterine PGF2alpha production is mediated, at least in part, by up-regulation of PLA2G6 expression and activity. In addition to its known inhibitory effect on oxytocin receptor expression, interferon-tau represses oxytocin-stimulated PLA2G6 expression and activity and this contributes to diminished PGF2alpha production. Furthermore, endometrial cell PGE2 biosynthesis was associated with PLA2G4C expression and activity and interferon-tau was stimulatory to this process.     

Multiplexed genotyping of ABC transporter polymorphisms with the Bioplex suspension array

We have developed and validated a consolidated bead-based genotyping platform, the Bioplex suspension array for simultaneous detection of multiple single nucleotide polymorphisms (SNPs) of the ATP-binding cassette transporters. Genetic polymorphisms have been known to influence therapeutic response and risk of disease pathologies. Genetic screening for therapeutic and diagnostic applications thus holds great promise in clinical management. The allele-specific primer extension (ASPE) reaction was used to assay 22 multiplexed SNPs for eight subjects. Comparison of the microsphere-based ASPE assay results to sequencing results showed complete concordance in genotype assignments. The Bioplex suspension array thus proves to be a reliable, cost-effective and high-throughput technological platform for genotyping. It can be easily adapted to customized SNP panels for specific applications involving large-scale mutation screening of clinically relevant markers.