Functionally Important Carboxyls in a Bacterial Homologue of the Vesicular Monoamine Transporter (VMAT)

Transporters essential for neurotransmission in mammalian organisms and bacterial multidrug transporters involved in antibiotic resistance are evolutionarily related. To understand in more detail the evolutionary aspects of the transformation of a bacterial multidrug transporter to a mammalian neurotransporter and to learn about mechanisms in a milieu amenable for structural and biochemical studies, we identified, cloned, and partially characterized bacterial homologues of the rat vesicular monoamine transporter (rVMAT2). We performed preliminary biochemical characterization of one of them, Brevibacillus brevis monoamine transporter (BbMAT), from the bacterium B. brevis. BbMAT shares substrates with rVMAT2 and transports them in exchange with >1H⁺, like the mammalian transporter. Here we present a homology model of BbMAT that has the standard major facilitator superfamily fold; that is, with two domains of six transmembrane helices each, related by 2-fold pseudosymmetry whose axis runs normal to the membrane and between the two halves. The model predicts that four carboxyl residues, a histidine, and an arginine are located in the transmembrane segments. We show here that two of the carboxyls are conserved, equivalent to the corresponding ones in rVMAT2, and are essential for H⁺-coupled transport. We conclude that BbMAT provides an excellent experimental paradigm for the study of its mammalian counterparts and bacterial multidrug transporters.     

Ecological niche modelling and population genetic analysis of Indian temperate bamboo Drepanostachyum falcatum in the western Himalayas

Journal of Plant Research - The present study was conducted to understand the key ecological and biological questions of conservation importance in Drepanostachyum falcatum which aimed to map...     

Regulatory enzymes of lipid metabolism in LA/N-cp rats

Hepatic activities of rate limiting enzymes in fatty acid and cholesterol synthesis and cholesterol degradation were determined in lean and obese LA/N-cp rats. The hepatic activities of acetyl-CoA carboxylase and fatty acid synthetase, the key enzymes of fatty acid synthesis and 3-hydroxy-3-methylglutaryl coenzyme A reductase (the rate limiting enzyme in cholesterol synthesis), were increased 2-fold in the obese rats as compared with their lean littermates. In contrast, the activity of cholesterol 7alpha-hydroxylase, the rate limiting enzyme of cholesterol degradation to bile acids, was significantly decreased by 28% in the obese group as compared with the control group. Significantly, compared with the control group, the obese animals exhibited similar magnitudes of differences in the activities of the above enzymes even when they were pair-fed with the control animals. Thus these differences in the obese group are not due to hyperphagia but possibly to hypersecretion of the lipogenic hormone, insulin in this strain. These results indicate that the LA/N-cp obese rat has twice the capacity to synthesize body fat and cholesterol but has a reduced capacity to degrade the cholesterol, leading to increased accumulation of cholesterol and fat.     

Glucose-Modified Low Density Lipoprotein Enhances Human Monocyte Chemotaxis

In diabetes mellitus the progression of atherosclerosis is accelerated. The interaction of glucose with athero-genic lipoproteins may be relevant to the mechanisms responsible for this vascular damage. The aim of this study was to examine the effect of glucose-modified low density lipoprotein (LDL) on human monocyte chemotaxis and to investigate the roles of oxidation and glycation in the generation of chemotactic LDL. Cu(II)-mediated LDL oxidation was potentiated by glucose in a dose-dependent manner and increased its chemotactic activity. Incubation with glucose alone, under conditions where very little oxidation was observed, also increased the chemotactic property of LDL. Neither diethylenetriamine pentaacetic acid (DETAPAC) nor aminoguanidine, which both inhibited LDL oxidation, completely inhibited the chemotactic activity of glycated oxidised LDL. The results suggest that both oxidation and glycation contribute to increased chemotactic activity.     

Chromosome 17 markers and von Recklinghausen neurofibromatosis: A genetic linkage study in a British population

A genetic linkage study of the RFLPs identified by nine DNA probes localized to the pericentromeric region and long arm of chromosome 17 has been undertaken in 16 families with von Recklinghausen neurofibromatosis (NF1). Close linkage has been shown with the markers CRI-L946 (D17S36), CRI-L581 (D17S37), p17H8 (D17Z1), and pA10-41 (D17S71). The ERBA1 and COL1A1 loci may also be closely linked, but the data are limited. The results for HOX2 and NGFR suggest only loose linkage with the NF1 gene, while no linkage was found between NF1 and the growth hormone locus. No suggestion of nonallelic heterogeneity of NF1 was found in this study.     

HORMONAL REQUIREMENTS OF EXCISED DIANTHUS CARYOPHYLLUS L. SHOOT APICAL MERISTEM IN VITRO

Whereas a medium containing kinetin alone enabled a few Dianthus caryophyllus L. apical meristem dome explants to develop into rooted plants, the highest frequency of plants was obtained in one containing supplements of both IAA and kinetin. In an unsupplemented medium, continued development required that explants have 2 pairs of primordial and a pair of expanding leaves. Kinetin alone caused production of many new leaves, but the development was significantly less than when it was furnished in combination with IAA. IAA given alone caused meristem explants to develop primarily callus, roots, and a few leaves. Gibberellin and abscisic acid were without promotive effects on leaf and shoot formation. A balance of hormonal substances, synthesized in young leaf structures and relocated to the meristem, is proposed as the fundamental mechanism that regulates new leaf initiation in the shoot apex.     

Organogenesis,embryogenesis, and synthetic seed production in <Emphasis Type="Italic">Arnebia euchroma</Emphasis>—A critically endangered medicinal plant of the Himalaya

Summary This is the first report of simultaneous organogenesis and somatic embryogenesis in Arnebia euchroma, a highly valued, critically endangered medicinal plant of the Himalaya. Root-derived callus showed only rhizogenesis, whereas leaf-derived callus showed simutaneous organogenesis and somatic embryogenesis. Organogenesis was optimal (12.2 shoots per culture) in 1 μM indole-3-butyric acid combined with 2.5 μM 6-benzyladenine and induction of somatic embryogenesis (16.3 embryos per culture) occurred in 2.5 μM indole-3-butyric acid combined with 2.5 μM 6-benzyladenine. Shoots rooted (100%) best in half-strength Murashige and Skoog (MS) medium supplemented with 2.0 μM indole-3-butyric acid. Early cotyledonary-stage embryos encapsulated with 3% sodium alginate and calcium nitrate (100 mM for 25 min) showed 60.6% germination in MS medium. Rooted shoots transferred to a mixture of sterile soil, sand, and peat (1∶1∶1 by volume) showed 72% survival ex vitro. Application of these protocols would be helpful in reducing pressure in natural populations, in genetic transformation studies, and in long-term storage of elite genotypes through synthetic seed production.     

GLUT4 expression and subcellular localization in the intrauterine growth-restricted adult rat female offspring

Intrauterine growth restriction (IUGR) leads to obesity, glucose intolerance, and type 2 diabetes mellitus in the adult. To determine the mechanism(s) behind this "metabolic imprinting" phenomenon, we examined the effect of total calorie restriction during mid- to late gestation modified by postnatal ad libitum access to nutrients (CM/SP) or nutrient restriction (SM/SP) vs. postnatal nutrient restriction alone (SM/CP) on skeletal muscle and white adipose tissue (WAT) insulin-responsive glucose transporter isoform (GLUT4) expression and insulin-responsive translocation. A decline in skeletal muscle GLUT4 expression and protein concentrations was noted only in the SM/SP and SM/CP groups. In contrast, WAT demonstrated no change in GLUT4 expression and protein concentrations in all experimental groups. The altered in utero hormonal/metabolic milieu was associated with a compensatory adaptation that persisted in the adult and consisted of an increase in the skeletal muscle basal plasma membrane-associated GLUT4 concentrations. This perturbation led to no further exogenous insulin-induced GLUT4 translocation, thereby disabling the insulin responsiveness of the skeletal muscle but retaining it in WAT. These changes, which present at birth, collectively maximize basal glucose transport to the compromised skeletal muscle with a relative resistance to exogenous/postprandial insulin. Preservation of insulin responsiveness in WAT may serve as a sink that absorbs postprandial nutrients that can no longer efficiently access skeletal muscle. We speculate that, in utero, GLUT4 aberrations may predict type 2 diabetes mellitus, whereas postnatal nutrient intake may predict obesity, thereby explaining the heterogeneous phenotype of the IUGR adult offspring.     

u-Genome: a database on genome design in unicellular genomes

Unicellular eukaryotes were among the first ones to be selected for complete genome sequencing because of the small size of their genomes and their interactions with humans and a broad range of animals and plants. Currently, ten completely sequenced unicellular genome sequences have been publicly released and as the number of available unicellular genomes increases, comparative genomics analysis within this group of organisms becomes more and more instructive. However, such an analysis is difficult to carry out without a suitable platform gathering not only the original annotations but also relevant information available in public databases or obtained by applying common bioinformatics methods. With the aim of solving these difficulties, we have developed a web-accessible database named u-Genome, the unicellular genome design database. The database is unique in featuring three datasets namely (1) orthologous proteins (2) paralogous proteins and (3) statistical distributions on exons, introns, intergenic DNA and correlations between them. A tool, Uniview, designed to visualize the gene structures for individual genes in the genome is also integrated. This database is of importance in understanding unicellular genome design and architecture and evolution related studies. The database is available through a web interface at http://sege.ntu.edu.sg/wester/ugenome.