Ionic interactions are essential for TRPV1 C-terminus binding to calmodulin

Calmodulin (CaM) is known to play an important role in the regulation of TRP channels activity. Although it has been reported that CaM binds to the C-terminus of TRPV1 (TRPV1-CT), no classic CaM-binding motif was found in this region. In this work, we explored this unusual TRPV1 CaM-binding motif in detail and found that five residues from a putative CaM-binding motif are important for TRPV1-CT’s binding to CaM, with arginine R785 being the most essential residue. The homology modelling suggests that a CaM-binding motif of TRPV1-CT forms an alpha helix that docks into the central cavity of CaM.     

Proteomic analysis uncovers a metabolic phenotype in C. elegans after nhr-40 reduction of function

Caenorhabditis elegans has an unexpectedly large number (284) of genes encoding nuclear hormone receptors, most of which are nematode-specific and are of unknown function. We have exploited comparative two-dimensional chromatography of synchronized cultures of wild type C. elegans larvae and a mutant in nhr-40 to determine if proteomic approaches will provide additional insight into gene function. Chromatofocusing, followed by reversed-phase chromatography and mass spectrometry, identified altered chromatographic patterns for a set of proteins, many of which function in muscle and metabolism. Prompted by the proteomic analysis, we find that the penetrance of the developmental phenotypes in the mutant is enhanced at low temperatures and by food restriction. The combination of our phenotypic and proteomic analysis strongly suggests that NHR-40 provides a link between metabolism and muscle development. Our results highlight the utility of comparative two-dimensional chromatography to provide a relatively rapid method to gain insight into gene function.     

A novel, cellulose synthesis inhibitory action of ancymidol impairs plant cell expansion

The co-ordination of cell wall synthesis with plant cell expansion is an important topic of contemporary plant biology research. In studies of cell wall synthesis pathways, cellulose synthesis inhibitors are broadly used. It is demonstrated here that ancymidol, known as a plant growth retardant primarily affecting gibberellin biosynthesis, is also capable of inhibiting cellulose synthesis. Its ability to inhibit cellulose synthesis is not related to its anti-gibberellin action and possesses some unique features never previously observed when conventional cellulose synthesis inhibitors were used. It is suggested that ancymidol targets the cell wall synthesis pathway at a regulatory step where cell wall synthesis and cell expansion are coupled. The elucidation of the ancymidol target in plant cells could potentially contribute to our understanding of cell wall synthesis and cell expansion control.     

Metachromatic leukodystrophy: magnetic resonance imaging (diffusion weighted image--DWI)

Ten years old patient with juvenile form of metachromatic leukodystrophy (MDL) is presented. Apart from standard magnetic resonance (MR) protocol, the diffusion weighted sequence was performed and apparent diffusion coefficient (ADC) map was constructed. The cause of restricted diffusion is considered to be the deposits of metachromatic substance in the intercellular space, apparently similar reason for decrease of speed of diffusion in other storage diseases.     

Induction of muscle thermogenesis by high-fat diet in mice: association with obesity-resistance

The obesogenic effect of a high-fat (HF) diet is counterbalanced by stimulation of energy expenditure and lipid oxidation in response to a meal. The aim of this study was to reveal whether muscle nonshivering thermogenesis could be stimulated by a HF diet, especially in obesity-resistant A/J compared with obesity-prone C57BL/6J (B/6J) mice. Experiments were performed on male mice born and maintained at 30 degrees C. Four-week-old mice were randomly weaned onto a low-fat (LF) or HF diet for 2 wk. In the A/J LF mice, cold exposure (4 degrees C) resulted in hypothermia, whereas the A/J HF, B/6J LF, and B/6J HF mice were cold tolerant. Cold sensitivity of the A/J LF mice was associated with a relatively low whole body energy expenditure under resting conditions, which was normalized by the HF diet. In both strains, the HF diet induced uncoupling protein-1-mediated thermogenesis, with a stronger induction in A/J mice. Only in A/J mice: 1) the HF diet augmented activation of whole body lipid oxidation by cold; and 2) at 30 degrees C, oxygen consumption, total content, and phosphorylation of AMP-activated protein kinase (AMPK), and AICAR-stimulated palmitate oxidation in soleus muscle was increased by the HF diet in parallel with significantly increased leptinemia. Gene expression data in soleus muscle of the A/J HF mice indicated a shift from carbohydrate to fatty acid oxidation. Our results suggest a role for muscle nonshivering thermogenesis and lipid oxidation in the obesity-resistant phenotype of A/J mice and indicate that a HF diet could induce thermogenesis in oxidative muscle, possibly via the leptin-AMPK axis.     

7H-Dibenzo[c,g]carbazole and 5,9-dimethyldibenzo[c,g]carbazole exert multiple toxic events contributing to tumor promotion in rat liver epithelial 'stem-like' cells

Immature liver progenitor cells have been suggested to be an important target of hepatotoxins and hepatocarcinogens. The goal of the present study was to assess the impact of 7H-dibenzo[c,g]carbazole (DBC) and its tissue-specific carcinogenic N-methyl (N-MeDBC) and 5,9-dimethyl (DiMeDBC) derivatives on rat liver epithelial WB-F344 cells, in vitro model of liver progenitor cells. We investigated the cellular events associated with both tumor initiation and promotion, such as activation of aryl hydrocarbon receptor (AhR), changes in expression of enzymes involved in metabolic activation of DBC and its derivatives, effects on cell cycle, cell proliferation/apoptosis and inhibition of gap junctional intercellular communication (GJIC). N-MeDBC, a tissue-specific sarcomagen, was only a weak inhibitor of GJIC or inducer of AhR-mediated activity, and it did not affect either cell proliferation or apoptosis. DBC was efficient GJIC inhibitor, while DiMeDBC manifested the strongest AhR inducing activity. Accordingly, DiMeDBC was also the most potent inducer of cytochrome P450 1A1 (CYP1A1) and CYP1A2 expression among the three compounds tested. Both DBC and DiMeDBC induced expression of CYP1B1 and aldo-keto reductase 1C9 (AKR1C9). N-MeDBC failed to significantly upregulate CYP1A1/2 and it only moderately increased CYP1B1 or AKR1C9. Only the potent liver carcinogens, DBC and DiMeDBC, caused a significant increase of p53 phosphorylation at Ser15, an increased accumulation of cells in S-phase and apoptosis at micromolar concentrations. In addition, DiMeDBC was found to stimulate cell proliferation of contact-inhibited WB-F344 cells at 1 microM concentration, which is a mode of action that might further contribute to its hepatocarcinogenicity. The present data seem to suggest that the AhR activation, induction of enzymes involved in metabolic activation, inhibition of GJIC or stimulation of cell proliferation might all contribute to the hepatocarcinogenic effects of DBC and DiMeDBC.     

Systemic immunization with an ALVAC-HIV-1/protein boost vaccine strategy protects rhesus macaques from CD4+ T-cell loss and reduces both systemic and mucosal simian-human immunodeficiency virus SHIVKU2 RNA levels

Transmission of human immunodeficiency virus type 1 (HIV-1) occurs primarily via the mucosal route, suggesting that HIV-1 vaccines may need to elicit mucosal immune responses. Here, we investigated the immunogenicity and relative efficacy of systemic immunization with two human ALVAC-HIV-1 recombinant vaccines expressing Gag, Pol, and gp120 (vCP250) or Gag, Pol, and gp160 (vCP1420) in a prime-boost protocol with their homologous vaccine native Env proteins. The relative efficacy was measured against a high-dose mucosal exposure to the pathogenic neutralization-resistant variant SHIV(KU2) (simian-human immunodeficiency virus). Systemic immunization with both vaccine regimens decreased viral load levels not only in blood but unexpectedly also in mucosal sites and protected macaques from peripheral CD4+ T-cell loss. This protective effect was stronger when the gp120 antigen was included in the vaccine. Inclusion of recombinant Tat protein in the boosting phase along with the Env protein did not contribute further to the preservation of CD4+ T cells. Thus, systemic immunization with ALVAC-HIV-1 vaccine candidates elicits anti-HIV-1 immune responses able to contain virus replication also at mucosal sites in macaques.     

Zinc(II) complexes with potent cyclin-dependent kinase inhibitors derived from 6-benzylaminopurine: synthesis, characterization, X-ray structures and biological activity

The synthesis, characterization and biological activity of the first zinc(II) complexes with potent inhibitors of cyclin-dependent kinases (CDKs) derived from 6-benzylaminopurine are described. Based on the results following from elemental analyses, infrared, NMR and ES+MS (electrospray mass spectra in the positive ion mode) spectroscopies, conductivity data, thermal analysis and X-ray structures, the tetrahedral Zn(II) complexes of the compositions [Zn(Olo)Cl(2)](n) (1), [Zn(iprOlo)Cl(2)](n) (2), [Zn(BohH(+))Cl(3)] x H(2)O (3) and [Zn(iprOloH(+))Cl(3)] x H(2)O (4) have been prepared, where Olo=2-(2-hydroxyethylamino)-6-benzylamino-9-methylpurine (Olomoucine), iprOlo=2-(2-hydroxyethylamino)-6-benzylamino-9-isopropylpurine (i-propyl-Olomoucine), Boh=2-(3-hydroxypropylamino)-6-benzylamino-9-isopropylpurine (Bohemine). The 1D-polymeric chain structure for [Zn(Olo)Cl(2)](n) (1) as well as the monomeric one for [Zn(BohH(+))Cl(3)] x H(2)O (3) and [Zn(iprOloH(+))Cl(3)] x H(2)O (4) have been revealed unambiguously by single crystal X-ray analyses. The 1D-polymeric chain of 1 consists of Zn(Olo)Cl(2) monomeric units in which the Zn(II) ion is coordinated by two chlorine atoms and one oxygen atom of the 2-hydroxyethylamino group of Olomoucine. The next monomeric unit is bonded to Zn(II) through the N7 atom of a purine ring. Thus, each of Zn(II) ions is tetrahedrally coordinated and a ZnCl(2)NO chromophore occurs in the complex 1. The complexes 3 and 4 are mononuclear species with a distorted tetrahedral arrangement of donor atoms around the Zn(II) ion with a ZnCl(3)N chromophore. The corresponding CDK inhibitor, i.e., both Boh and iprOlo, is coordinated to Zn(II) via the N7 atom of the purine ring in 3 and 4. The cytotoxicity of the zinc(II) complexes against human melanoma, sarcoma, leukaemia and carcinoma cell lines has been determined as well as the inhibition of the CDK2/cyclin E kinase. A relationship between the structure and biological activity of the complexes is also discussed.