Quantitative Mass Density Image Reconstructed from the Complex X-Ray Refractive Index

We demonstrate a new analytical X-ray computed tomography technique for visualizing and quantifying the mass density of materials comprised of low atomic number elements with unknown atomic ratios. The mass density was obtained from the experimentally observed ratio of the imaginary and real parts of the complex X-ray refractive index. An empirical linear relationship between the X-ray mass attenuation coefficient of the materials and X-ray energy was found for X-ray energies between 8 keV and 30 keV. The mass density image of two polymer fibers was quantified using the proposed technique using a scanning-type X-ray microbeam computed tomography system equipped with a wedge absorber. The reconstructed mass density agrees well with the calculated one.     

Occurrence of a novel strain of Scirtothrips dorsalis (Thysanoptera: Thripidae) in Japan and development of its molecular diagnostics

Scirtothrips dorsalis Hood is a cosmopolitan and polyphagous thrips species. Recently, a novel strain of S. dorsalis attacking capsicum crops was found in Japan. A molecular phylogenetic analysis using mitochondrial cytochrome c oxidase subunit I sequences revealed that the capsicum-associated populations were genetically different from Japanese native strains and were closely related to Southeast Asian populations. We named the capsicum-associated populations “strain C” and the Japanese native ones “strain YT”. A total of 10 haplotypes were found in strain C and 26 in strain YT. To differentiate the two strains, we developed a multiplex-PCR method using the ribosomal ITS2 region.     

A conserved α helix of Bcs1, a mitochondrial AAA chaperone,is required for the Respiratory Complex III maturation

Bcs1 is a transmembrane chaperone in the mitochondrial inner membrane, and is required for the mitochondrial Respiratory Chain Complex III assembly. It has been shown that the highly-conserved C-terminal region of Bcs1 including the AAA ATPase domain in the matrix side is essential for the chaperone function. Here we describe the importance of the N-terminal short segment located in the intermembrane space in the Bcs1 function. Among the N-terminal 44 amino acid residues of yeast Bcs1, the first 37 residues are dispensable whereas a hydrophobic amino acid in the residue 38 is essential for integration of Rieske Iron-sulfur Protein into the premature Complex III from the mitochondrial matrix. Substitution of the residue 38 by a hydrophilic amino acid residue affects conformation of Bcs1 and interactions with other proteins. The evolutionarily-conserved short α helix of Bcs1 in the intermembrane space is an essential element for the chaperone function.     

Activation of activin type IB receptor signals in pancreatic β cells leads to defective insulin secretion through the attenuation of ATP-sensitive K channel activity

In studies of gene-ablated mice, activin signaling through activin type IIB receptors (ActRIIB) and Smad2 has been shown to regulate not only pancreatic β cell mass but also insulin secretion. However, it still remains unclear whether gain of function of activin signaling is involved in the modulation of pancreatic β cell mass and insulin secretion. To identify distinct roles of activin signaling in pancreatic β cells, the Cre-loxP system was used to activate signaling through activin type IB receptor (ActRIB) in pancreatic β cells. The resultant mice (pancreatic β cell-specific ActRIB transgenic (Tg) mice; ActRIBCAβTg) exhibited a defect in glucose-stimulated insulin secretion (GSIS) and a progressive impairment of glucose tolerance. Patch-clamp techniques revealed that the activity of ATP-sensitive K⁺ channels (K_ATP channels) was decreased in mutant β cells. These results indicate that an appropriate level of activin signaling may be required for GSIS in pancreatic β cells, and that activin signaling involves modulation of K_ATP channel activity.     

Effects of type III antifreeze protein on sperm and embryo cryopreservation in rabbit

We investigated the effects of antifreeze protein (AFP) III supplementation on the cryopreservation of rabbit sperm cells and embryos. Ejaculated semen was collected from male Japanese white (JW) rabbits and divided into four AFP-supplemented groups (0.1 μg/ml, 1 μg/ml, 10 μg/ml, 100 μg/ml) and one control group with no AFP-supplementation. The semen samples were treated with egg-yolk HEPES extender containing 6% acetamide before the sperm was cooled from room temperature to 5 °C, then packed into sperm straws. The straws were frozen in steam of liquid nitrogen (LN₂) and then preserved in the LN₂. The motility of the sperm after thawing in 37 °C water was analyzed. The percentage of rapidly motile sperm in the 1 μg/ml AFP group was significantly higher than in the control group. Morulae were collected from female JW rabbits and divided into three AFP-supplemented groups (100 ng/ml, 500 ng/ml, 1000 ng/ml) and one control group. The morulae, immersed in an embryo-freezing solution (M199-HEPES containing 20% ethylene glycol, 20% dimethylsulfoxide, 10% fetal bovine serum and 0.25 M sucrose), were packed into open pulled embryo straws and vitrified in LN₂. The frozen embryos were thawed in the embryo-freezing solution, and the rates of embryo survival and development to blastocyte stage were analyzed after incubation for 72 h. The development rate of the embryos in the 500 ng/ml AFP group was significantly higher than in the control group, but that in the 1000 ng/ml AFP group was significantly lower. In conclusion, the appropriate dose of AFP III increased the number of rapidly motile sperm and embryo survival following freezing and thawing. The results suggest that supplementation with AFP III can increase the efficiency of cryopreservation of rabbit sperm cells and embryos.     

Replacement of C305 in Heart/Muscle-Type Isozyme of Human Carnitine Palmitoyltransferase I with Aspartic Acid and Other Amino Acids

Liver- and heart/muscle-type isozymes of human carnitine palmitoyltransferase I (L- and M-CPTI, respectively) show a certain similarity in their amino acid sequences, and mutation studies on the conserved amino acids between these two isozymes often show essentially the same effects on their enzymatic properties. Earlier mutation studies on C305 in human M-CPTI and its counterpart residue, C304, in human L-CPTI showed distinct effects of the mutations, especially in the aspect of enzyme stability; however, simple comparison of these effects on the conserved Cys residue between L- and M-CPTI was difficult, because these studies were carried out using different expression systems and distinct amino acids as replacements. In the present study, we carried out mutation studies on the C305 in human M-CPTI using COS cells for the expression system. Our results showed that C305 was replaceable with aspartic acid but that substitution with other amino acids caused both loss of function and reduced expression.     

Zierane sesquiterpene lactone,cembrane and fusicoccane diterpenoids,from the Tahitian liverwort Chandonanthus hirtellus

Cembrane-type diterpenoids, 13,18,20-epi-iso-chandonanthone (1) and (8E)-4α-acetoxy-12α,13α-epoxycembra-1(15),8-diene (2), two fusicoccane-type diterpenoids, fusicoauritone 6α-methyl ether (3) and 6β,10β-epoxy-5β-hydroxyfusicocc-2-ene (4) and a zierane sesquiterpene γ-lactone, chandolide (5) were isolated from the Tahitian liverwort Chandonanthus hirtellus (Web.) Mitt., together with eight known diterpenoids, chandonanthine (6), fusicogigantone A (7), fusicogigantone B (8), fusicogigantepoxide (9), anadensin (10), fusicoauritone (11), ent-verticillol (12) and ent-epi-verticillol (13). Their structures were established by a combination of extensive NMR spectroscopy and/or X-ray crystallographic analyses. Compounds 1, 5 and 10 showed weak cytotoxic activity against HL-60. Compound 3 also indicated weak cytotoxic activity against KB cell lines.     

Molecular basis for defect in Alix-binding by alternatively spliced isoform of ALG-2 (ALG-2<Superscript>ΔGF122</Superscript>) and structural roles of F122 in target recognition

Background  

ALG-2 (a gene product of PDCD6) belongs to the penta-EF-hand (PEF) protein family and Ca²⁺-dependently interacts with various intracellular proteins including mammalian Alix, an adaptor protein in the ESCRT system. Our previous X-ray crystal structural analyses revealed that binding of Ca²⁺ to EF3 enables the side chain of R125 to move enough to make a primary hydrophobic pocket (Pocket 1) accessible to a short fragment of Alix. The side chain of F122, facing a secondary hydrophobic pocket (Pocket 2), interacts with the Alix peptide. An alternatively spliced shorter isoform, designated ALG-2^ΔGF122, lacks Gly¹²¹Phe¹²² and does not bind Alix, but the structural basis of the incompetence has remained to be elucidated.