Augmented Reticular Thalamic Bursting and Seizures in Scn1a-Dravet Syndrome

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Babesia bovis: effects of cysteine protease inhibitors on in vitro growth

In the present study, we examined the effects of four kinds of cysteine protease inhibitors (E64, E64d, leupeptin, and ALLN) on the in vitro asexual growth of Babesia bovis. Of these, only the lipophilic inhibitors, E64d and ALLN, were found to effectively inhibit the growth of B. bovis. In further experiments, E64d, but not ALLN, significantly suppressed the parasite’s invasion of host erythrocytes, while both chemicals, especially ALLN, inhibited the parasite’s replication within the infected erythrocytes. These data suggested the presence of cysteine protease(s) derived from B. bovis, in which the protease(s) would play important roles in the erythrocyte invasion and/or replication processes of the parasite.     

Valproate induces apoptosis by inducing accumulation of neutral lipids which was prevented by disruption of the SIR2 gene in Saccharomyces cerevisiae

We investigated the participation of HDACs in VPA induced apoptosis in Saccharomyces cerevisiae. VPA (20 mM) induced apoptosis in several HDAC mutants, including PRD3 and HDA1-disrupted cells and SIR2 over expressing cells, as well as in wild-type cells but not SIR2-disrupted cells. Intracellular reactive oxygen species and neutral lipid content increased markedly in all kinds of HDAC mutant cells tested except for SIR2-disrupted cells. Thus, these results suggest that 20 mM VPA induces neutral lipid accumulation and apoptosis-like features in S. cerevisiae, and that VPA-induced apoptosis was evaded by deletion of SIR2.     

Genomewide association analysis in diverse inbred mice: power and population structure

The discovery of quantitative trait loci (QTL) in model organisms has relied heavily on the ability to perform controlled breeding to generate genotypic and phenotypic diversity. Recently, we and others have demonstrated the use of an existing set of diverse inbred mice (referred to here as the mouse diversity panel, MDP) as a QTL mapping population. The use of the MDP population has many advantages relative to traditional F(2) mapping populations, including increased phenotypic diversity, a higher recombination frequency, and the ability to collect genotype and phenotype data in community databases. However, these methods are complicated by population structure inherent in the MDP and the lack of an analytical framework to assess statistical power. To address these issues, we measured gene expression levels in hypothalamus across the MDP. We then mapped these phenotypes as quantitative traits with our association algorithm, resulting in a large set of expression QTL (eQTL). We utilized these eQTL, and specifically cis-eQTL, to develop a novel nonparametric method for association analysis in structured populations like the MDP. These eQTL data confirmed that the MDP is a suitable mapping population for QTL discovery and that eQTL results can serve as a gold standard for relative measures of statistical power.     

A neural-specific F-box protein Fbs1 functions as a chaperone suppressing glycoprotein aggregation

Fbs1 is an F-box protein present abundantly in the nervous system. Similar to the ubiquitously expressed Fbs2, Fbs1 recognizes N-glycans at the innermost position as a signal for unfolded glycoproteins, probably in the endoplasmic reticulum-associated degradation pathway. Here, we show that the in vivo majority of Fbs1 is present as Fbs1-Skp1 heterodimers or Fbs1 monomers but not SCF(Fbs1) complex. The inefficient SCF complex formation of Fbs1 and the restricted presence of SCF(Fbs1) bound on the endoplasmic reticulum membrane were due to the short linker sequence between the F-box domain and the sugar-binding domain. In vitro, Fbs1 prevented the aggregation of the glycoprotein through the N-terminal unique sequence of Fbs1. Our results suggest that Fbs1 assists clearance of aberrant glycoproteins in neuronal cells by suppressing aggregates formation, independent of ubiquitin ligase activity, and thus functions as a unique chaperone for those proteins.     

Serotonin differentially regulates short- and long-term prediction of rewards in the ventral and dorsal striatum

Background

The ability to select an action by considering both delays and amount of reward outcome is critical for maximizing long-term benefits. Although previous animal experiments on impulsivity have suggested a role of serotonin in behaviors requiring prediction of delayed rewards, the underlying neural mechanism is unclear.

Methodology/Principal Findings

To elucidate the role of serotonin in the evaluation of delayed rewards, we performed a functional brain imaging experiment in which subjects chose small-immediate or large-delayed liquid rewards under dietary regulation of tryptophan, a precursor of serotonin. A model-based analysis revealed that the activity of the ventral part of the striatum was correlated with reward prediction at shorter time scales, and this correlated activity was stronger at low serotonin levels. By contrast, the activity of the dorsal part of the striatum was correlated with reward prediction at longer time scales, and this correlated activity was stronger at high serotonin levels.

Conclusions/Significance

Our results suggest that serotonin controls the time scale of reward prediction by differentially regulating activities within the striatum.     

Induction of increase in intracellular calcium concentration of embryonic cells and acceleration of morphogenetic cell movements during amphibian gastrulation by a 50-Hz magnetic field

The influence of an alternating electromagnetic field (EMF) on early development of amphibian embryos was examined. When the embryos developed under the influence of a low-frequency EMF (50 Hz, 5-30 mT), the rate of early development was accelerated. The effect of EMF was exerted preferentially at the gastrula stage, and the period of gastrulation was shortened. Histological observations showed that EMF promoted morphogenetic cell movements during the gastrulation. The concentration of intracellular free Ca2+ ([Ca2+]i) in the embryonic cells under the influence of EMF was analyzed using Fura-2, an indicator of the intracellular concentration of calcium ions. The influence of EMF on [Ca2+]i was analyzed in embryonic cells isolated from blastula, gastrula, and neurula, EMF increased a [Ca2+]i particularly in the cells isolated from gastrula. Our results suggest that EMF specifically increased the [Ca2+]i of gastrula cells, thereby, accelerating the rate of morphogenetic cell movements during gastrulation.     

Early development in utero of bovine nuclear transfer embryos using early G1 and G0 phase cells

Bovine somatic cell nuclear transfer (NT) embryos can develop to normal calves, but the success rates are still quite low. Recently, enhanced development of bovine NT embryos to full term has been achieved using fibroblasts at the early G1 phase instead of cells at the quiescent (G0) phase. In the present study, we examined the morphological development in utero of NT embryos using early G1 phase cells (eG1-NT embryos) and G0 phase cells (G0-NT embryos). We produced eG1- and G0-NT blastocysts, and then they were transferred to recipient heifers for transient development in utero up to day 14 of gestation. In vitro-fertilized (IVF), parthenogenetic and artificially inseminated (AI) embryos were used as controls. The rate of formation of embryonic disks of the recovered embryos was the same among the groups of eG1-NT, IVF, and AI embryos (p>0.05). The formation rate in eG1-NT embryos was significantly higher than that in G0-NT embryos (p<0.05). The lengths of eG1-NT embryos were the same as those of IVF, parthenogenetic, and AI embryos (p>0.05), but significantly shorter than those of G0-NT embryos (p<0.01). We conclude that the morphological development of day 14 embryos derived from eG1-NT embryos was mostly similar to that of AI embryos, but that the morphological development of G0-NT embryos was abnormally large and different from that of AI and eG1-NT embryos.     

Rapid SNP diagnostics using asymmetric isothermal amplification and a new mismatch-suppression technology

We developed a rapid single nucleotide polymorphism (SNP) detection system named smart amplification process version 2 (SMAP 2). Because DNA amplification only occurred with a perfect primer match, amplification alone was sufficient to identify the target allele. To achieve the requisite fidelity to support this claim, we used two new and complementary approaches to suppress exponential background DNA amplification that resulted from mispriming events. SMAP 2 is isothermal and achieved SNP detection from whole human blood in 30 min when performed with a new DNA polymerase that was cloned and isolated from Alicyclobacillus acidocaldarius (Aac pol). Furthermore, to assist the scientific community in configuring SMAP 2 assays, we developed software specific for SMAP 2 primer design. With these new tools, a high-precision and rapid DNA amplification technology becomes available to aid in pharmacogenomic research and molecular-diagnostics applications.     

Direct interactions between NEDD8 and ubiquitin E2 conjugating enzymes upregulate cullin-based E3 ligase activity

Although cullin-1 neddylation is crucial for the activation of SCF ubiquitin E3 ligases, the underlying mechanisms for NEDD8-mediated activation of SCF remain unclear. Here we demonstrate by NMR and mutational studies that NEDD8 binds the ubiquitin E2 (UBC4), but not NEDD8 E2 (UBC12). Our data imply that NEDD8 forms an active platform on the SCF complex for selective recruitment of ubiquitin-charged E2s in collaboration with RBX1, and thereby upregulates the E3 activity.