Monoamine oxidase A gene polymorphisms and enzyme activity associated with risk of gout in Taiwan aborigines

Taiwanese aborigines have a high prevalence of hyperuricemia and gout. Uric acid levels and urate excretion have correlated with dopamine-induced glomerular filtration response. MAOs represent one of the major renal dopamine metabolic pathways. We aimed to identify the monoamine oxidase A (MAOA, Xp11.3) gene variants and MAO-A enzyme activity associated with gout risk. This study was to investigate the association between gout and the MAOA single-nucleotide polymorphisms (SNPs) rs5953210, rs2283725, and rs1137070 as well as between gout and the COMT SNPs rs4680 Val158Met for 374 gout cases and 604 controls. MAO-A activity was also measured. All three MAOA SNPs were significantly associated with gout. A synonymous MAOA SNP, rs1137070 Asp470Asp, located in exon 14, was associated with the risk of having gout (P = 4.0 × 10^?5, adjusted odds ratio 1.46, 95% confidence intervals [CI]: 1.11–1.91). We also showed that, when compared to individuals with the MAOA GAT haplotype, carriers of the AGC haplotype had a 1.67-fold (95% CI: 1.28–2.17) higher risk of gout. Moreover, we found that MAOA enzyme activity correlated positively with hyperuricemia and gout (P for trend = 2.00 × 10^?3 vs. normal control). We also found that MAOA enzyme activity by rs1137070 allele was associated with hyperuricemia and gout (P for trend = 1.53 × 10^?6 vs. wild-type allele). Thus, our results show that some MAOA alleles, which have a higher enzyme activity, predispose to the development of gout.     

A novel definition of the multivariate coefficient of variation

The coefficient of variation CV (%) is widely used to measure the relative variation of a random variable to its mean or to assess and compare the performance of analytical techniques/equipments. A review is made of the existing multivariate extensions of the univariate CV where, instead of a random variable, a random vector is considered, and a novel definition is proposed. The multivariate CV obtained only requires the calculation of the mean vector, the covariance matrix and simple quadratic forms. No matrix inversion is needed which makes the new approach equally attractive in high dimensional as in very small sample size problems. As an illustration, the method is applied to electrophoresis data from external quality assessment in laboratory medicine, to phenotypic characteristics of pocket gophers and to a microarray data set.     

First record of Cercidiphylloxylon(Cercidiphyllaceae)from the Palaeocene of Fushun,NE China

Cercidiphylloxylon spenceri(Brett)Pearson is described from the Lizigou Formation,Palaeocene in China.The growth rings are distinct; pores are diffuse,solitary,with somewhat angular outlines in cross section;vessel elements long with long scalariform perforation plates; intervessel pitting is opposite to scalariform; fibertracheids are present; axial parenchyma is scarce; rays are mostly biseriate and heterogeneous.All wood characters of the fossil specimen fall into the range of those of extant Cercidiphyllum(Cercidiphyllaceae).The finding is one of the earliest fossil wood records of Cercidiphyllaceae.     

Synthesis,binding and bioactivity of γ-methylene γ-lactam ecdysone receptor ligands: Advantages of QSAR models for flexible receptors

Nuclear hormone receptors, such as the ecdysone receptor, often display a large amount of induced fit to ligands. The size and shape of the binding pocket in the EcR subunit changes markedly on ligand binding, making modelling methods such as docking extremely challenging. It is, however, possible to generate excellent 3D QSAR models for a given type of ligand, suggesting that the receptor adopts a relatively restricted number of binding site configurations or ‘attractors’. We describe the synthesis, in vitro binding and selected in vivo toxicity data for γ-methylene γ-lactams, a new class of high-affinity ligands for ecdysone receptors from Bovicola ovis (Phthiraptera) and Lucilia cuprina (Diptera). The results of a 3D QSAR study of the binding of methylene lactams to recombinant ecdysone receptor protein suggest that this class of ligands is indeed recognised by a single conformation of the EcR binding pocket.     

Introduction of Virulence Markers in PB2 of Pandemic Swine-Origin Influenza Virus Does Not Result in Enhanced Virulence or Transmission

In the first 6 months of the H1N1 swine-origin influenza virus (S-OIV) pandemic, the vast majority of infections were relatively mild. It has been postulated that mutations in the viral genome could result in more virulent viruses, leading to a more severe pandemic. Mutations E627K and D701N in the PB2 protein have previously been identified as determinants of avian and pandemic influenza virus virulence in mammals. These mutations were absent in S-OIVs detected early in the 2009 pandemic. Here, using reverse genetics, mutations E627K, D701N, and E677G were introduced into the prototype S-OIV A/Netherlands/602/2009, and their effects on virus replication, virulence, and transmission were investigated. Mutations E627K and D701N caused increased reporter gene expression driven by the S-OIV polymerase complex. None of the three mutations affected virus replication in vitro. The mutations had no major impact on virus replication in the respiratory tracts of mice and ferrets or on pathogenesis. All three mutant viruses were transmitted via aerosols or respiratory droplets in ferrets. Thus, the impact of key known virulence markers in PB2 in the context of current S-OIVs was surprisingly small. This study does not exclude the possibility of emergence of S-OIVs with other virulence-associated mutations in the future. We conclude that surveillance studies aimed at detecting S-OIVs with increased virulence or transmission should not rely solely on virulence markers identified in the past but should include detailed characterization of virus phenotypes, guided by genetic signatures of viruses detected in severe cases of disease in humans.The new H1N1 swine-origin influenza virus (S-OIV) recently emerged to cause the first influenza pandemic in 40 years (2). The S-OIV presumably emerged from pigs, as its genome was shown to consist of six gene segments of “triple-reassortant” swine viruses and two of “Eurasian lineage” swine viruses (9). The start of the S-OIV pandemic has been relatively mild, with a clinical spectrum ranging from mild upper respiratory tract illness to sporadic cases of severe pneumonia leading to acute respiratory distress syndrome (22). As of 15 November 2009, worldwide, more than 206 countries have reported laboratory-confirmed cases of S-OIV infection, including over 6,770 deaths (32).In previous influenza pandemics, such as the Spanish influenza pandemic of 1918 and the Hong Kong influenza pandemic of 1968, a first wave of cases of relatively mild illnesses was followed by more severe subsequent waves (29). The reason for this increased severity has remained largely unknown, but one possible explanation could be that the pandemic viruses required further adaptation to the human host, resulting in the emergence of viruses that were more virulent than those of the first wave. Such adaptive changes could occur by gene reassortment between cocirculating influenza A viruses or by mutation.In the past decade, determinants of influenza A virus virulence have been mapped using reverse genetics with a variety of pandemic, epidemic, and zoonotic influenza viruses. Mutations affecting virulence and host range have frequently been mapped to hemagglutinin (HA) and neuraminidase (NA) in relation to their interaction with sialic acids, the virus receptors on host cells (11, 18, 30). Nonstructural protein 1 (NS1) has been implicated in the virulence of highly pathogenic avian influenza (HPAI) virus H5N1 and the 1918 H1N1 virus, as the NS1 proteins of these viruses were shown to act as strong antagonists of the interferon pathways (10, 25). Furthermore, the polymerase genes, in particular the PB2 gene, have been shown to be important determinants of virulence in the HPAI H5N1 and H7N7 viruses and of transmission in the 1918 H1N1 virus (11, 21, 31). One of the most commonly identified virulence markers to date is E627K in PB2. The glutamic acid (E) residue is found generally in avian influenza viruses, while human viruses have a lysine (K), and this mutation has been described as a determinant of the host range in vitro (28). When avian viruses lacking the E627K substitution were passaged in mice, the viruses acquired the mutation spontaneously upon a single passage (15, 17). In the HPAI H5N1 and H7N7 viruses, E627K was shown to be the prime determinant of pathogenesis in mice (11, 21, 23). Given that all human and many zoonotic influenza viruses of the last century contained 627K (1), it was surprising that the S-OIV had 627E.Additionally, the aspartate (D)-to-asparagine (N) mutation at position 701 of PB2, which was shown to compensate for the absence of E627K, has also not been detected in S-OIV (27). This D701N mutation has previously been shown to expand the host range of avian H5N1 to mice and humans (3, 15) and to increase virus transmission in guinea pigs (27). Thus, S-OIV was the first known human pandemic virus with 627E and 701D, and it has been speculated that S-OIV could mutate into a more virulent form by acquiring one of these mutations, or both.On 8 May 2009, the detection of another mutation in the PB2 gene of S-OIV, an E-to-glycine (G) mutation at position 667, was reported (http://www.promedmail.org/pls/apex/f?p=2400:1000, archive no. 20090508.1722). It has previously been suggested that the E667G substitution in PB2 of HPAI H5N1 virus was under positive selection and possibly played a role in sustainable transmission in humans (14).On 28 September 2009, detection of the E627K mutation in PB2 of S-OIVs of two individuals in the Netherlands was reported (http://www.promedmail.org/pls/apex/f?p=2400:1000, archive no. 20090928.3394) and raised concern about the possible enhanced replication of the S-OIV in humans, possibly associated with increased virulence. To date, the D701N mutation in PB2 has not been reported in any of the S-OIVs sequenced, and additional viruses with mutation E627K have not been recorded, either. In contrast, viruses with E677G have been reported from the United States, Canada, Germany, the United Kingdom, Norway, and France, according to the public sequence databases.Here, the effects of the E627K, D701N, and E677G mutations in the PB2 genes of S-OIVs was investigated using genetically engineered influenza viruses based on a prototype S-OIV, A/Netherlands/602/2009. Polymerase activity was measured in minigenome assays in human 293T cells, virus replication was analyzed in Madin-Darby Canine kidney (MDCK) cells, virulence was tested in mouse and ferret models, and transmission by aerosols or respiratory droplets was tested in ferrets. In contrast to the earlier assumptions based on experience with other influenza A viruses, S-OIVs with E627K, D701N, or E677G in PB2 did not show a marked increase in virulence or transmission compared to the wild-type virus.     

Structure of a Eukaryotic Nonribosomal Peptide Synthetase Adenylation Domain That Activates a Large Hydroxamate Amino Acid in Siderophore Biosynthesis

Nonribosomal peptide synthetases (NRPSs) are large, multidomain proteins that are involved in the biosynthesis of an array of secondary metabolites. We report the structure of the third adenylation domain from the siderophore-synthesizing NRPS, SidN, from the endophytic fungus Neotyphodium lolii. This is the first structure of a eukaryotic NRPS domain, and it reveals a large binding pocket required to accommodate the unusual amino acid substrate, N^δ-cis-anhydromevalonyl-N^δ-hydroxy-l-ornithine (cis-AMHO). The specific activation of cis-AMHO was confirmed biochemically, and an AMHO moiety was unambiguously identified as a component of the fungal siderophore using mass spectroscopy. The protein structure shows that the substrate binding pocket is defined by 17 amino acid residues, in contrast to both prokaryotic adenylation domains and to previous predictions based on modeling. Existing substrate prediction methods for NRPS adenylation domains fail for domains from eukaryotes due to the divergence of their signature sequences from those of prokaryotes. Thus, this new structure will provide a basis for improving prediction methods for eukaryotic NRPS enzymes that play important and diverse roles in the biology of fungi.     

TNFAIP8: A new effector for Galpha(i) coupling to reduce cell death and induce cell transformation

Galpha(i)‐coupled receptors comprise a diverse family of receptors that induce transformation by largely unknown mechanisms. We previously found that the Galpha(i)‐coupled dopamine‐D2short (D2S) receptor transforms Balb‐D2S cells via Gαi3. To identify new Gαi effectors, a yeast two‐hybrid screen was done using constitutively active Gαi3‐Q204L as bait, and tumor necrosis factor‐alpha (TNFα)‐induced protein 8 (TNFAIP8, SCC‐S2/NDED/GG2‐1) was identified. In contrast, TNFAIP8‐related TIPE1 and TIPE2 showed a very weak interaction with Gαi3. In yeast mating, in vitro pull‐down, co‐immunoprecipitation and bioluminescence resonance energy transfer (BRET) assays, TNFAIP8 preferentially interacted with activated Gαi proteins, consistent with direct Gαi‐TNFAIP8 coupling. Over‐expression or depletion of TNFAIP8 using antisense constructs in Balb‐D2S cells did not affect D2S‐induced signaling to Gαi‐dependent inhibition of cAMP. In contrast, antisense depletion of TNFAIP8 completely inhibited spontaneous and D2S‐induced foci formation, consistent with a role for TNFAIP8 in Gαi‐dependent transformation. To address possible mechanisms, the effect of D2S signaling via TNFAIP8 on TNFα action was examined. D2S receptor activation inhibited TNFα‐induced cell death in Balb‐D2S cells, but not in cells depleted of TNFAIP8. However, depletion of TNFAIP8 did not prevent D2S‐induced inhibition of TNFα‐mediated caspase activation, suggesting that D2S/TNFAIP8‐induced protection from TNFα‐induced cell death is caspase‐independent. The data suggest that Gαi‐TNFAIP8‐mediated rescue of pre‐oncogenic cells enhances progression to oncogenic transformation, providing a selective target to inhibit cellular transformation. J. Cell. Physiol. 225: 865–874, 2010. © 2010 Wiley‐Liss, Inc.     

Distribution of Small Integrin-binding Ligand, N-linked Glycoproteins (SIBLING) in the Articular Cartilage of the Rat Femoral Head

The small integrin-binding ligand, N-linked glycoprotein (SIBLING) family is closely related to osteogenesis. Until recently, little was known about their existence in articular cartilage. In this study, we systematically evaluated the presence and distribution of four SIBLING family members in rat femoral head cartilage: dentin matrix protein 1 (DMP1), bone sialoprotein (BSP), osteopontin (OPN), and dentin sialophosphoprotein (DSPP). First, non-collagenous proteins were extracted and then separated by ion-exchange chromatography. Next, the protein extracts eluted by chromatography were analyzed by Stains-all staining and Western immunoblotting. IHC was used to assess the distribution of these four SIBLING family members in the femoral head cartilage. Both approaches showed that all the four SIBLING family members are expressed in the femoral head cartilage. IHC showed that SIBLING members are distributed in various locations throughout the articular cartilage. The NH₂-terminal fragments of DMP1, BSP, and OPN are present in the cells and in the extracellular matrix, whereas the COOH-terminal fragment of DMP1 and the NH₂-terminal fragment of DSPP are primarily intracellularly localized in the chondrocytes. The presence of the SIBLING family members in the rat femoral head cartilage suggests that they may play important roles in chondrogenesis. (J Histochem Cytochem 58:1033–1043, 2010)     

Competition between wild-type and a marked recombinant baculovirus (Spodoptera exigua nucleopolyhedrovirus) with enhanced speed of action in insect larvae

Competition between virus genotypes in insect hosts is a key element of virus fitness, affecting their long-term persistence in agro-ecosystems. Little information is available on virus competition in insect hosts or during serial passages from one cohort of hosts to the next. Here we report on the competition between two genotypes of Spodoptera exigua nucleopolyhedrovirus (SeMNPV), when serially passaged as mixtures in cohorts of 4th instar S. exigua larvae. One of the genotypes was a SeMNPV wild-type isolate, SeUS1, while the other was a SeMNPV recombinant (SeMNPV-XD1) having a greater speed of kill than SeUS1. SeXD1 lacks a suite of genes, including the ecdysteroid UDP-glucosyl transferase (egt) gene. SeXD1 expresses the green fluorescent protein (GFP) gene, enabling the identification of SeXD1 in cell culture and in insects. The relative proportion of SeUS1 and SeXD1 in successive passages of mixed infections in various ratios was determined by plaque assays of budded virus from infected larvae and by polymerase chain reactions and restriction enzyme analyses. The SeUS1 genotype outcompeted recombinant SeXD1 over successive passages. Depending on the initial virus genotype ratio, the recombinant SeXD1 was no longer detected after 6-12 passages. A mathematical model was developed to characterize the competition dynamics. Overall, the ratio SeUS1/XD1 increased by a factor 1.9 per passage. The findings suggest that under the experimental conditions recombinant SeXD1 is displaced by the wild-type strain SeUS1, but further studies are needed to ascertain that this is also the case when the same baculoviruses would be used in agro-ecosystems.