Role of FK506-binding protein 12 in development of the chick embryonic heart

A cDNA encoding chicken FK506-binding protein 12 (FKBP12) was isolated and sequenced. The predicted amino acid sequence of the chicken protein shows high homology to those of FKBP12 proteins of other species ranging from human to frog. The possible role of FKBP12 in chick embryonic cardiac development was examined. Northern blot analysis revealed that FKBP12 mRNA is distributed widely in chick embryos, being especially abundant in the heart; the amount of FKBP12 mRNA in the embryonic heart decreased with time. Administration of FK506 to chick embryos at 7 to 9 days resulted in marked cardiac enlargement. FK506 also reduced the expression of myosin, induced a more elongated cell morphology, and impaired network formation in cultured chick embryonic cardiomyocytes. These results suggest that FKBP12 is important in the regulation of contractile function and phenotypic expression in chick cardiomyocytes during embryonic development.     

Identification of immunostimulatory DNA-induced genes by suppression subtractive hybridization

Bacterial DNA and related synthetic immunostimulatory oligodeoxyribo-nucleotides (ISS-ODN) have stimulatory effects on mammalian immune cells through a Toll-like receptor, TLR9. Genes upregulated in ISS-ODN-stimulated immune cells are obviously significant to delineate the mechanism of the induced innate immunity. Employing suppression subtractive hybridization (SSH), we have generated a profile of genes induced by ISS-ODN in spleen cells. Sequencing of 87 clones isolated by the SSH showed 39 clones corresponding to known mouse genes in the public database. Eleven clones appeared to possess 80-90% homology with known mouse genes and the remaining 37 clones showed no significant homology with any known mouse genes. A series of known genes which have not previously been reported to be induced with ISS-ODN were confirmed to be induced in ISS-ODN-stimulated bone marrow-derived macrophages: NF-kappaB p105, IRF-1, PA28beta, IRG2, and MyD88. These genes were suggested to be involved in the molecular process of innate host defense mechanisms.     

Effects of glucose, vitamins, and DO concentrations on pyruvate fermentation using Torulopsis glabrata IFO 0005 with metabolic flux analysis

The effects of glucose, vitamins, and DO concentrations on efficient pyruvic acid fermentation were investigated using Torulopsis glabrata IFO 0005, and a novel biphasic culture method was developed on the basis of the metabolic flux analysis. T. glabrata requires the four vitamins nicotinic acid (NA), thiamine hydrochloride (B(1)), pyridoxine hydrochloride, and biotin for cell growth. The deficiency of these vitamins plays an essential role in pyruvate fermentation. In the present study, we considered the effects of the first two vitamins on the pyruvate fermentation. On the basis of several batch and fed-batch experiments, it was found that, as a result of glucose inhibition of cell growth, the initial glucose concentration should be around 30-40 g/L, and the glucose concentration during fermentation should be controlled at high level around 30 g/L. On the basis of an analysis of carbon flux distribution, a biphasic fermentation method was developed where the cultivation started with a high DO (at 40-50% of air saturation) for efficient cell growth and then was reduced to 5-10% for efficient pyruvate production. Since a fair amount of ethanol was formed when the DO concentration was decreased, the addition of NA turned out to be effective in reducing the ethanol formation. This may be due to a relaxing of the requirement for NADH oxidation by the alcohol dehydrogenase pathway. Since B(1) affects both the pyruvate dehydrogenase complex and pyruvate decarboxylase, its initial concentration must be carefully determined by considering both the cell growth and pyruvate production phases.     

Comparison in gene expression of secretory human endometrium using laser microdissection

Background  

The endometrium prepares for implantation under the control of steroid hormones. It has been suggested that there are complicated interactions between the epithelium and stroma in the endometrium during menstrual cycle. In this study, we demonstrate a difference in gene expression between the epithelial and stromal areas of the secretory human endometrium using microdissection and macroarray technique.     

Crystal structure of outer surface protein C (OspC) from the Lyme disease spirochete, Borrelia burgdorferi

Outer surface protein C (OspC) is a major antigen on the surface of the Lyme disease spirochete, Borrelia burgdorferi, when it is being transmitted to humans. Crystal structures of OspC have been determined for strains HB19 and B31 to 1.8 and 2.5 A resolution, respectively. The three-dimensional structure is predominantly helical. This is in contrast to the structure of OspA, a major surface protein mainly present when spirochetes are residing in the midgut of unfed ticks, which is mostly beta-sheet. The surface of OspC that would project away from the spirochete's membrane has a region of strong negative electrostatic potential which may be involved in binding to positively charged host ligands. This feature is present only on OspCs from strains known to cause invasive human disease.     

Interaction of binuclear xylylthiolato(2,2',2"-terpyridine)platinum(II) complexes with DNA

The new binuclear platinum(II) complexes, (1,3-benzenedimethanethiolate-S)di(2,2',2"-terpyridine)diplatinum(II) chloride tetrahydrate, 5, and (1,4-benzenedimethanethiolate-S)di(2,2',2"-terpyridine)diplatinum(II) chloride tetrahydrate, 6, were synthesized in order to investigate the binding of platinum(II) complex with calf thymus DNA, which was examined by UV and CD spectroscopies. Complex 5 interacted strongly with DNA by intercalation compared to 6.     

Multistep fluorescence resonance energy transfer in sequential chromophore array constructed on oligo-DNA assemblies

Sequential arrays of chromophores at regulated distances were constructed on a noncovalent DNA molecular assembly system in aqueous media. Photoinduced fluorescence resonance energy transfer (FRET) behaviors were then observed. We designed a number of chromophore/oligo-DNA conjugates with varying sequences. The chromophores eosin (Eo), TexasRed (TR), and tetramethylrhodamine (Rho) were employed as the energy donor, acceptor, and mediator, respectively, based on overlapping excitation and emission spectra. The chromophores were attached via aminolinkers to the 5'-terminals of 10mer oligo-DNAs consisting of AT rich sequences. The arrangement of Eo-Rho or Rho-TR with 10-residue (1 pitch of duplex) distances was ensured by duplex formation of the conjugates with a 20mer matrix oligo-DNA composed of complementary sequences to the conjugates. Single-step FRET from Eo to Rho and from Rho to TR was confirmed on the duplex. The three chromophore conjugates were then mixed with longer matrix oligo-DNAs (30 or 40mer) consisting of complementary sequences to the conjugates, producing Eo-(Rho)(n)-TR (n = 1 or 2) arrays with 10-residue distances. Multistep FRET from Eo to TR through the Rho mediator(s) was observed on the molecular assemblies. This photoenergy transmission system offers a good model for a photoenergy transmission system mimicking photosynthetic systems.