Protein Interaction between Ameloblastin and Proteasome Subunit α Type 3 Can Facilitate Redistribution of Ameloblastin Domains within Forming Enamel

Enamel is a bioceramic tissue composed of thousands of hydroxyapatite crystallites aligned in parallel within boundaries fabricated by a single ameloblast cell. Enamel is the hardest tissue in the vertebrate body; however, it starts development as a self-organizing assembly of matrix proteins that control crystallite habit. Here, we examine ameloblastin, a protein that is initially distributed uniformly across the cell boundary but redistributes to the lateral margins of the extracellular matrix following secretion thus producing cell-defined boundaries within the matrix and the mineral phase. The yeast two-hybrid assay identified that proteasome subunit α type 3 (Psma3) interacts with ameloblastin. Confocal microscopy confirmed Psma3 co-distribution with ameloblastin at the ameloblast secretory end piece. Co-immunoprecipitation assay of mouse ameloblast cell lysates with either ameloblastin or Psma3 antibody identified each reciprocal protein partner. Protein engineering demonstrated that only the ameloblastin C terminus interacts with Psma3. We show that 20S proteasome digestion of ameloblastin in vitro generates an N-terminal cleavage fragment consistent with the in vivo pattern of ameloblastin distribution. These findings suggest a novel pathway participating in control of protein distribution within the extracellular space that serves to regulate the protein-mineral interactions essential to biomineralization.     

Molecular dynamics simulation on the decomposition of type SII hydrogen hydrate and the performance of tetrahydrofuran as a stabiliser

Molecular dynamics simulation is used to study the decomposition and stability of SII hydrogen and hydrogen/tetrahydrofuran (THF) hydrates at 150 K, 220 K and 100 bar. The modelling of the microscopic decomposition process of hydrogen hydrate indicates that the decomposition of hydrogen hydrate is led by the diffusive behaviour of H₂ molecules. The hydrogen/THF hydrate presents higher stability, by comparing the distributions of the tetrahedral angle of H₂O molecules, radial distribution functions of H₂O molecules and mean square displacements or diffusion coefficients of H₂O and H₂ molecules in hydrogen hydrate with those in hydrogen/THF hydrate. It is also found that the resistance of the diffusion behaviour of H₂O and H₂ molecules can be enhanced by encaging THF molecules in the (5¹²6⁴) cavities. Additionally, the motion of THF molecules is restricted due to its high interaction energy barrier. Accordingly, THF, as a stabiliser, is helpful in increasing the stability of hydrogen hydrate.     

New cationic lipids form channel-like pores in phospholipid bilayers

Two representatives of a new class of cationic lipids were found to have high pore-forming activity in planar bilayer membranes. These molecules, called BHHD-TADC and BHTD-TADC, have qualitatively similar effects on phospholipid membranes. Addition of 2.5-5 micro M of either of them to the membrane bathing solutions resulted in formation of long-lived anion-selective pores with conductance in the range 0.1-2 nS in 0.1 M KCl. Pore formation was found to be dependent on the potential applied to the membrane. When negative potential was applied to membrane at the side of addition, the rate of pore formation was much lower compared to when the positive potential was applied. Dependence of pore formation on compound concentration was highly nonlinear, indicating that this process requires assembly of molecules in the membrane. Addition of any of these compounds on both sides of the membrane increased the efficiency of pore formation by one to two orders of magnitude. Pore formation was strongly pH dependent. Although pores were formed with high efficiency at pH 6.5, only occasional fluctuations of membrane conductance were observed at pH 7.5. Possible mechanisms of new compounds biological activity are discussed.     

Synchronization regulation in a model of coupled neural masses

A model of coupled neural masses can generate seizure-like events and dynamics similar to those observed during interictal to ictal transitions and thus can be used for theoretical study of the control of epileptic seizures. In an effort to understand the mechanisms underlying epileptic seizures and how to avoid them, we added a control input to this model. Epileptic seizures are always accompanied by hypersynchronous firing of neurons, so research on synchronization among cortical areas is significant for seizure control. In this study, principal component analysis (PCA) was used to identify synchronization clusters composed of several neural masses. A method for calculating the synchronization cluster strength and participation rate is presented. The synchronization cluster strength can be used to identify synchronization clusters and the participation rate can be employed to identify neural masses that participate in the clusters. Each synchronization cluster is controlled as a whole using a proportional-integral-derivative (PID) controller. We illustrate these points using coupled neural mass models of synchronization to show their responses to increased (between node) coupling with and without control. Experiment results indicated that PID control can effectively regulate synchronization between neural masses and has the potential for seizure prevention.     

High-performance single-chip exon capture allows accurate whole exome sequencing using the Illumina Genome Analyzer

Here we present an adaptation of NimbleGen 2.1M-probe array sequence capture for whole exome sequencing using the Illumina Genome Analyzer (GA) platform. The protocol involves two-stage library construction. The specificity of exome enrichment was approximately 80% with 95.6% even coverage of the 34 Mb target region at an average sequencing depth of 33-fold. Comparison of our results with whole genome shot-gun resequencing results showed that the exome SNP calls gave only 0.97% false positive and 6.27% false negative variants. Our protocol is also well suited for use with whole genome amplified DNA. The results presented here indicate that there is a promising future for large-scale population genomics and medical studies using a whole exome sequencing approach.     

Efficient production of <Emphasis Type="Italic">Agrobacterium rhizogenes</Emphasis>-transformed roots and composite plants in peanut (<Emphasis Type="Italic">Arachis hypogaea</Emphasis> L.)

Recalcitrance of most large-seeded legumes, such as peanut, to regeneration and genetic transformation has hampered studies on gene function and efforts for genetic improvement. Agrobacterium rhizogenes-mediated transformation provides a system for rapid and efficient transformation of plant tissues. In this study, embryonic axes along with cotyledons of peanut were injected with a suspension culture of A. rhizogenes using microliter syringes. The influence of several factors such as plant genotype, A. rhizogenes culture stage, co-culture period of A. rhizogenes, and acetosyringone concentration in the co-cultivation medium have been evaluated. It is found that A. rhizogenes-mediated transformation of peanut is genotype-independent. Up to 61% transformation was recorded when embryonic axes were co-cultivated with 5 × 10⁷ A. rhizogenes cells from logarithmic phase for 2 days on co-culture medium containing 50 μmol l⁻¹ acetosyringone. Composite plants with transgenic roots were harvested after 45 days of treatment. Furthermore, this method was applied to assess the insecticidal activity of a synthetic cry8Ea1 gene against Holotrichia parallela in transgenic roots of peanut.