Cellular distribution of Hsp70 expression in rat skeletal muscles. Effects of moderate exercise training and chronic hypoxia

Rat hindlimb muscles constitutively express the inducible heat shock protein 72 (Hsp70), apparently in proportion to the slow myosin content. Since it remains controversial whether chronic Hsp70 expression reflects the overimposed stress, we investigated Hsp70 cellular distribution in fast muscles of the posterior rat hindlimb after (1) mild exercise training (up to 30 m/min treadmill run for 1 h/day), which induces a remodeling in fast fiber composition, or (2) prolonged exposure to normobaric hypoxia (10%O₂), which does not affect fiber-type composition. Both conditions increased significantly protein Hsp70 levels in the skeletal muscle. Immunohistochemistry showed the labeling for Hsp70 in subsets of both slow/type 1 and fast/type 2A myofibers of control, sedentary, and normoxic rats. Endurance training increased about threefold the percentage of Hsp70-positive myofibers (P<0.001), and changed the distribution of Hsp70 immunoreactivity, which involved a larger subset of both type 2A and intermediate type 2A/2X myofibers (P<0.001) and vascular smooth muscle cells. Hypoxia induced Hsp70 immunoreactivity in smooth muscle cells of veins and did not increase the percentage of Hsp70-positive myofibers; however, sustained exposure to hypoxia affected the distribution of Hsp70 immunoreactivity, which appeared detectable in a very small subset of type 2A fibers, whereas it concentrated in type 1 myofibers (P<0.05) together with the labeling for heme-oxygenase isoform 1, a marker of oxidative stress. Therefore, the chronic induction of Hsp70 expression in rat skeletal muscles is not obligatory related to the slow fiber phenotype but reveals the occurrence of a stress response.     

Molecular Characterization of Three Small Isometric-Headed Bacteriophages Which Vary in Their Sensitivity to the Lactococcal Phage Resistance Plasmid pTR2030

Lactococcus lactis LMA12-4 is a pTR2030 transconjugant that has been used as an industrial starter culture because of its resistance to phages predominant in cheese plants. Plasmid pTR2030 interferes with susceptible phages in this host strain via two mechanisms, restriction and modification (R/M) and abortive infection (Hsp). After prolonged use of LMA12-4 transconjugants in the industry, two different bacteriophages, designated nck202.48 (48) and nck202.50 (50), were isolated which could produce plaques on LMA12-4 containing pTR2030. In this study, these two phages were characterized and compared with a third phage, nck202.31 (31), which is susceptible to both the R/M and Hsp activities encoded by pTR2030. Phage 48 was not susceptible to inhibition by Hsp, whereas 50 was unaffected by either the R/M or Hsp mechanisms. All three were small isometric-headed phages, but small differences were noted between the phages in the structural details of the tail base plate, susceptibility to chloroform treatment, and requirements for calcium infectivity. The phage genomes were all between 29.9 and 31.9 kb in length. Phages 31 and 48 harbored cohesive ends, whereas the phage 50 genome was circularly permuted, terminally redundant, and carried a putative packaging initiation site. DNA-DNA hybridization experiments conducted between the phages revealed a common region in 48 and 50 that may correlate with the resistance of the two phages to the Hsp-abortive infection induced by pTR2030. Phage 50 also harbored DNA sequences that shared homology to pTR2030 in the region where R/M activities have been localized on the plasmid. Molecular characterization of the three phages localized regions within the genomes of the pTR2030-resistant phages that may be responsible for circumventing plasmid-encoded Hsp and R/M defense mechanisms in lactococci.     

Isolation and Characterization of Five Erwinia amylovora Bacteriophages and Assessment of Phage Resistance in Strains of Erwinia amylovora

Phages able to infect the fire blight pathogen Erwinia amylovora were isolated from apple, pear, and raspberry tissues and from soil samples collected at sites displaying fire blight symptoms. Among a collection of 50 phage isolates, 5 distinct phages, including relatives of the previously described phages Ea1 and Ea7 and 3 novel phages named Ea100, Ea125, and Ea116C, were identified based on differences in genome size and restriction fragment pattern. Ea1, the phage distributed most widely, had an approximately 46-kb genome which exhibited some restriction site variability between isolates. Phages Ea100, Ea7, and Ea125 each had genomes of approximately 35 kb and could be distinguished by their EcoRI restriction fragment patterns. Ea116C contained an approximately 75-kb genome. Ea1, Ea7, Ea100, Ea125, and Ea116C were able to infect 39, 36, 16, 20, and 40, respectively, of 40 E. amylovora strains isolated from apple orchards in Michigan and 8, 12, 10, 10, and 12, respectively, of 12 E. amylovora strains isolated from raspberry fields (Rubus spp.) in Michigan. Only 22 of 52 strains were sensitive to all five phages, and 23 strains exhibited resistance to more than one phage. Ea116C was more effective than the other phages at lysing E. amylovora strain Ea110 in liquid culture, reducing the final titer of Ea110 by >95% when added at a ratio of 1 PFU per 10 CFU and by 58 to 90% at 1 PFU per 10⁵ CFU.     

Admixture Mapping of 15,280 African Americans Identifies Obesity Susceptibility Loci on Chromosomes 5 and X

The prevalence of obesity (body mass index (BMI) ≥30 kg/m²) is higher in African Americans than in European Americans, even after adjustment for socioeconomic factors, suggesting that genetic factors may explain some of the difference. To identify genetic loci influencing BMI, we carried out a pooled analysis of genome-wide admixture mapping scans in 15,280 African Americans from 14 epidemiologic studies. Samples were genotyped at a median of 1,411 ancestry-informative markers. After adjusting for age, sex, and study, BMI was analyzed both as a dichotomized (top 20% versus bottom 20%) and a continuous trait. We found that a higher percentage of European ancestry was significantly correlated with lower BMI (ρ=−0.042, P=1.6×10⁻⁷). In the dichotomized analysis, we detected two loci on chromosome X as associated with increased African ancestry: the first at Xq25 (locus-specific LOD=5.94; genome-wide score=3.22; case-control Z=−3.94); and the second at Xq13.1 (locus-specific LOD=2.22; case-control Z=−4.62). Quantitative analysis identified a third locus at 5q13.3 where higher BMI was highly significantly associated with greater European ancestry (locus-specific LOD=6.27; genome-wide score=3.46). Further mapping studies with dense sets of markers will be necessary to identify the alleles in these regions of chromosomes X and 5 that may be associated with variation in BMI.     

Specificity of the STAT4 genetic association for severe disease manifestations of systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is a genetically complex disease with heterogeneous clinical manifestations. A polymorphism in the STAT4 gene has recently been established as a risk factor for SLE, but the relationship with specific SLE subphenotypes has not been studied. We studied 137 SNPs in the STAT4 region genotyped in 4 independent SLE case series (total n=1398) and 2560 healthy controls, along with clinical data for the cases. Using conditional testing, we confirmed the most significant STAT4 haplotype for SLE risk. We then studied a SNP marking this haplotype for association with specific SLE subphenotypes, including autoantibody production, nephritis, arthritis, mucocutaneous manifestations, and age at diagnosis. To prevent possible type-I errors from population stratification, we reanalyzed the data using a subset of subjects determined to be most homogeneous based on principal components analysis of genome-wide data. We confirmed that four SNPs in very high LD (r²=0.94 to 0.99) were most strongly associated with SLE, and there was no compelling evidence for additional SLE risk loci in the STAT4 region. SNP rs7574865 marking this haplotype had a minor allele frequency (MAF)=31.1% in SLE cases compared with 22.5% in controls (OR=1.56, p=10⁻¹⁶). This SNP was more strongly associated with SLE characterized by double-stranded DNA autoantibodies (MAF=35.1%, OR=1.86, p<10⁻¹⁹), nephritis (MAF=34.3%, OR=1.80, p<10⁻¹¹), and age at diagnosis<30 years (MAF=33.8%, OR=1.77, p<10⁻¹³). An association with severe nephritis was even more striking (MAF=39.2%, OR=2.35, p<10⁻⁴ in the homogeneous subset of subjects). In contrast, STAT4 was less strongly associated with oral ulcers, a manifestation associated with milder disease. We conclude that this common polymorphism of STAT4 contributes to the phenotypic heterogeneity of SLE, predisposing specifically to more severe disease.     

Neuropeptide Y Gene Polymorphisms Confer Risk of Early-Onset Atherosclerosis

Neuropeptide Y (NPY) is a strong candidate gene for coronary artery disease (CAD). We have previously identified genetic linkage to familial CAD in the genomic region of NPY. We performed follow-up genetic, biostatistical, and functional analysis of NPY in early-onset CAD. In familial CAD (GENECARD, N=420 families), we found increased microsatellite linkage to chromosome 7p14 (OSA LOD=4.2, p=0.004) in 97 earliest age-of-onset families. Tagged NPY SNPs demonstrated linkage to CAD of a 6-SNP block (LOD=1.58–2.72), family-based association of this block with CAD (p=0.02), and stronger linkage to CAD in the earliest age-of-onset families. Association of this 6-SNP block with CAD was validated in: (a) 556 non-familial early-onset CAD cases and 256 controls (OR 1.46–1.65, p=0.01–0.05), showing stronger association in youngest cases (OR 1.84–2.20, p=0.0004–0.09); and (b) GENECARD probands versus non-familial controls (OR 1.79–2.06, p=0.003–0.02). A promoter SNP (rs16147) within this 6-SNP block was associated with higher plasma NPY levels (p=0.04). To assess a causal role of NPY in atherosclerosis, we applied the NPY1-receptor–antagonist BIBP-3226 adventitially to endothelium-denuded carotid arteries of apolipoprotein E-deficient mice; treatment reduced atherosclerotic neointimal area by 50% (p=0.03). Thus, NPY variants associate with atherosclerosis in two independent datasets (with strong age-of-onset effects) and show allele-specific expression with NPY levels, while NPY receptor antagonism reduces atherosclerosis in mice. We conclude that NPY contributes to atherosclerosis pathogenesis.     

Prophage Origin of a Virulent Phage Appearing on Fermentations of Lactobacillus casei S-1

For protection from the abnormal fermentation of Lactobacillus casei S-1 caused by contamination of a virulent phage, FSV, the origin of this phage was studied. Morphologies, viral structural proteins, and DNA structures of three independent isolates of FSV were compared with those of FSW, which is lysogenized in strain S-1. The results showed (i) that the morphology of FSV phages is indistinguishable from that of FSW and (ii) that all viral structural components found in FSW are present in the particles of FSV's. In addition, restriction endonuclease analyses of viral DNA showed that the HindIII-digested fragments of FSW DNA, the sum of which covered at least 94.7% of this phage genome, were conserved in the FSV DNA digests. Results of Southern filter hybridization of the S-1 and prophage-cured cell (C239) DNAs with FSV DNA as a probe revealed that C239 had lost most of the FSV DNA sequence, whereas S-1 had about one copy of the FSV DNA sequence. These results indicate that virulent phage FSV is derived from the lysogenized phage FSW. Therefore, the appearance of FSV can be eliminated by using the prophage-cured derivative of S-1.     

Restricted Diffusion in Biophysical Systems: Experiment

The pulsed-gradient spin echo nuclear magnetic resonance (PGSENMR) technique was used to measure restricted diffusion of water in three types of animal tissue: human blood plasma and red cells; rat and rabbit heart; rat and rabbit liver. Characteristic lengths (L) for restriction of diffusion are estimated from dependence on the measuring time. Limitations on the range of observable restrictive lengths (1.5-15 μm) are discussed.

The decrease in diffusivity due to 1 μm alumina powder (volume fraction = 0.18) in glycerin/water mixtures agrees with the Wang theory assuming spherical particles and no hydration. The characteristic length (L 4 μm) is larger than the particle size (1 μm) or separation (1.8 μm). Comparison of the diffusivities in tissues at short diffusion times with the Wang theory indicates some bound or trapped water.

For packed red blood cells, a restriction (L 2.3 μm) was attributed tothe red cell membrane. A permeability p 0.014 cm/s may be estimated from the decrease in diffusivity. Average values of diffusivity ratio in heart were: 0.36 ± 0.02 for rat; and 0.26 ± 0.03 for rabbit; and in liver: 0.25 ± 0.01 for rat; 0.25 ± .04 for 10-day old rabbit; and 0.195 ± 0.03 for 2-yr old rabbit. A restriction (L 2.7 μm) in rat liver probably results from the mitochondria.

     

Effect of turgor pressure and cell size on the wall elasticity of plant cells

Direct measurements of the volumetric elastic modulus, , of cells of a higher plant were performed on the epidermal bladder cells of Mesembryanthemum crystallinum using a pressure probe technique. Measurements on giant algal cells (Valonia, Nitellopsis) are given for comparison. Giant celled algae and M. crystallinum bladders have elastic moduli, , which depend strongly on turgor pressure, P, and on cell volume, V. The values of Mesembryanthemum bladders range between 5 bar at zero pressure and 100 bar at full turgor pressure (3-4 bar). increased with cell size (volume) at a given turgor pressure, and this volume dependence was pronounced more in the high pressure range. From the (P) characteristics, complete volume-pressure curves were obtained for Mesembryanthemum bladders and giant algal cells. The results suggest that the (P) and (V) characteristics of all plant cells are similar. The significance of the pressure and volume effects for the water relations and growth processes of plant cells is discussed briefly.     

MDM2 interacts with the C-terminus of the catalytic subunit of DNA polymerase epsilon

MDM2 is induced by p53 in response to cellular insults such as DNA damage and can have effects upon the cell cycle that are independent or downstream of p53. We used a yeast two-hybrid screen to identify proteins that bind to MDM2 and which therefore might be involved in these effects. We found that MDM2 can bind to the C-terminus of the catalytic subunit of DNA polymerase (DNA pol ), to a region that is known to be essential in yeast. In an in vitro system we confirmed that MDM2 could bind to the homologous regions of both mouse and human DNA pol and to full-length human DNA pol . DNA pol co-immunoprecipitated with MDM2 from transfected H1299 cells and also from a HeLa cell nuclear extract. We show here that the DNA pol -interacting domain of MDM2 is located between amino acids 50 and 166. Our studies provide evidence that MDM2 interacts with a region of DNA pol that plays a critical role in the function of DNA pol .     

Interaction between Light Quality and Light Quantity in the Photoregulation of Anthocyanin Production

The interaction between phytochrome photoequilibrium () and photon flux in the photoregulation of anthocyanin production under prolonged irradiation was studied in seedlings of Brassica oleracea L. and Lycopersicon esculentum Mill. In cabbage, anthocyanin production increases with decreasing , reaching a maximum at the lowest value ( = 0.13) used in this study; in tomato, the extent of the response is higher at intermediate values, reaching a maximum at = 0.46. In cabbage, the response increases with increasing photon flux at all values; however, the response to changes in photon flux is minimal at = 0.85, and, at = 0.13, minimal at photon fluxes higher than 5 micromolar per square meter per second. In tomato, the response increases with increasing photon flux at = 0.46, 0.65, and 0.85, the response to changes in photon fluxes being minimal at = 0.85; at = 0.13 and 0.29 the response first increases (significantly at = 0.29 and minimally at = 0.13) and then decreases with increasing photon fluxes, the transition occurring at about 1 micromolar per square meter per second at = 0.13, and at 5 micromolar per square meter per second at = 0.29. The patterns of light quality-quantity interaction in the photoregulation of anthocyanin production are significantly different in cabbage and tomato and are also significantly different than those observed for other photomorphogenic responses to prolonged irradiations.     

Saturation Mutagenesis of Toluene ortho-Monooxygenase of Burkholderia cepacia G4 for Enhanced 1-Naphthol Synthesis and Chloroform Degradation

Directed evolution of toluene ortho-monooxygenase (TOM) of Burkholderia cepacia G4 previously created the hydroxylase α-subunit (TomA3) V106A variant (TOM-Green) with increased activity for both trichloroethylene degradation (twofold enhancement) and naphthalene oxidation (six-times-higher activity). In the present study, saturation mutagenesis was performed at position A106 with Escherichia coli TG1/pBS(Kan)TOMV106A to improve TOM activity for both chloroform degradation and naphthalene oxidation. Whole cells expressing the A106E variant had two times better naphthalene-to-1-naphthol activity than the wild-type cells (V_max of 9.3 versus 4.5 nmol·min⁻¹·mg of protein⁻¹ and unchanged K_m), and the regiospecificity of the A106E variant was unchanged, with 98% 1-naphthol formed, as was confirmed with high-pressure liquid chromatography. The A106E variant degrades its natural substrate toluene 63% faster than wild-type TOM does (2.12 ± 0.07 versus 1.30 ± 0.06 nmol·min⁻¹·mg of protein⁻¹ [mean ± standard deviation]) at 91 μM and has a substantial decrease in regiospecificity, since o-cresol (50%), m-cresol (25%), and p-cresol (25%) are formed, in contrast to the 98% o-cresol formed by wild-type TOM. The A106E variant also has an elevated expression level compared to that of wild-type TOM, as evidenced by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Another variant, the A106F variant, has 2.8-times-better chloroform degradation activity based on gas chromatography (V_max of 2.61 versus 0.95 nmol·min⁻¹·mg of protein⁻¹ and unchanged K_m) and chloride release (0.034 ± 0.002 versus 0.012 ± 0.001 nmol·min⁻¹·mg of protein⁻¹). The A106F variant also was expressed at levels similar to those of wild-type TOM and 62%-better toluene oxidation activity than wild-type TOM (2.11 ± 0.3 versus 1.30 ± 0.06 nmol·min⁻¹·mg of protein⁻¹). A shift in regiospecificity of toluene hydroxylation was also observed for the A106F variant, with o-cresol (28%), m-cresol (18%), and p-cresol (54%) being formed. Statistical analysis was used to estimate that 292 colonies must be screened for a 99% probability that all 64 codons were sampled during saturation mutagenesis.     

Pasteurella pestis Bacteriophage H and Escherichia coli Bacteriophage φII Are Nearly Identical

Electron microscopy examination of II-H deoxyribonucleic acid heteroduplexes, together with polyacrylamide gel electrophoretic analyses of phage ribonucleic acid species and proteins labeled in II or H-infected cells, demonstrates that Pasteurella pestis phage H is nearly identical to coliphage II.     

Differential functional behavior of viral phi29, Nf and GA-1 SSB proteins

DNA replication of 29 and related phages takes place via a strand displacement mechanism, a process that generates large amounts of single-stranded DNA (ssDNA). Consequently, phage-encoded ssDNA-binding proteins (SSBs) are essential proteins during phage 29-like DNA replication. In the present work we analyze the helix-destabilizing activity of the SSBs of 29 and the related phages Nf and GA-1, their ability to eliminate non-productive binding of 29 DNA polymerase to ssDNA and their stimulatory effect on replication by 29 DNA polymerase in primed M13 ssDNA replication, a situation that resembles type II replicative intermediates that occur during 29-like DNA replication. Significant differences have been appreciated in the functional behavior of the three SSBs. First, the GA-1 SSB is able to display helix-destabilizing activity and to stimulate dNTP incorporation by 29 DNA polymerase in the M13 DNA replication assay, even at SSB concentrations at which the 29 and Nf SSBs do not show any effect. On the other hand, the 29 SSB is the only one of the three SSBs able to increase the replication rate of 29 DNA polymerase in primed M13 ssDNA replication. From the fact that the 29 SSB, but not the Nf SSB, stimulates the replication rate of Nf DNA polymerase we conclude that the different behaviors of the SSBs on stimulation of the replication rate of 29 and Nf DNA polymerases is most likely due to formation of different nucleoprotein complexes of the SSBs with the ssDNA rather than to a specific interaction between the SSB and the corresponding DNA polymerase. A model that correlates the thermodynamic parameters that define SSB–ssDNA nucleoprotein complex formation with the functional stimulatory effect of the SSB on 29-like DNA replication has been proposed.     

Progress in the discovery of selective, high affinity A2B adenosine receptor antagonists as clinical candidates

The selective, high affinity A_2B adenosine receptor (AdoR) antagonists that were synthesized by several research groups should aid in determining the role of the A_2B AdoR in inflammatory diseases like asthma or rheumatoid arthritis (RA) and angiogenic diseases like diabetic retinopathy or cancer. CV Therapeutics scientists discovered the selective, high affinity A_2B AdoR antagonist 10, a 8-(4-pyrazolyl)-xanthine derivative [CVT-6883, K_i(hA_2B)=22 nM; K_i(hA₁)=1,940 nM; K_i(hA_2A)=3,280; and K_i(hA₃)=1,070 nM] that has favorable pharmacokinetic (PK) properties (t_1/2=4 h and F>35% rat). Compound 10 demonstrated functional antagonism at the A_2B AdoR (K_B=6 nM) and efficacy in a mouse model of asthma. In two phase 1 clinical trials, CVT-6883 was found to be safe, well tolerated, and suitable for once daily dosing. A second compound 20, 8-(5-pyrazolyl)-xanthine, has been nominated for development from Baraldi’s group in conjunction with King Pharmaceuticals that has favorable A_2B AdoR affinity and selectivity [K_i(hA_2B)=5.5 nM; K_i(hA₁) >1,000 nM; K_i(hA_2A) >1,000; and K_i(hA₃) >1,000 nM], and it has been demonstrated to be a functional antagonist. A third compound 32, a 2-aminopyrimidine, from the Almirall group has high A_2B AdoR affinity and selectivity [K_i(hA_2B)=17 nM; K_i(hA₁) >1,000 nM; K_i(hA_2A) >2,500; and K_i(hA₃) >1,000 nM], and 32 has been moved into preclinical safety testing. Since three highly selective, high affinity A_2B AdoR antagonists have been nominated for development with 10 (CVT-6883) being the furthest along in the development process, the role of the A_2B AdoR in various disease states will soon be established.     

Exocyclic amino groups of flanking guanines govern sequence-dependent adduct conformations and local structural distortions for minor groove-aligned benzo[a]pyrenyl-guanine lesions in a GG mutation hotspot context

The environmental carcinogen benzo[a]pyrene (BP) is metabolized to reactive diol epoxides that bind to cellular DNA by predominantly forming N²-guanine adducts (G*). Mutation hotspots for these adducts are frequently found in 5′-···GG··· dinucleotide sequences, but their origins are poorly understood. Here we used high resolution NMR and molecular dynamics simulations to investigate differences in G* adduct conformations in 5′-···CG*GC··· and 5′-···CGG*C··· sequence contexts in otherwise identical 12-mer duplexes. The BP rings are positioned 5′ along the modified strand in the minor groove in both cases. However, subtle orientational differences cause strong distinctions in structural distortions of the DNA duplexes, because the exocyclic amino groups of flanking guanines on both strands compete for space with the BP rings in the minor groove, acting as guideposts for placement of the BP. In the 5′-···CGG*C··· case, the 5′-flanking G · C base pair is severely untwisted, concomitant with a bend deduced from electrophoretic mobility. In the 5′-···CG*GC··· context, there is no untwisting, but there is significant destabilization of the 5′-flanking Watson–Crick base pair. The minor groove width opens near the lesion in both cases, but more for 5′-···CGG*C···. Differential sequence-dependent removal rates of this lesion result and may contribute to the mutation hotspot phenomenon.     

Meta-Analysis of 28,141 Individuals Identifies Common Variants within Five New Loci That Influence Uric Acid Concentrations

Elevated serum uric acid levels cause gout and are a risk factor for cardiovascular disease and diabetes. To investigate the polygenetic basis of serum uric acid levels, we conducted a meta-analysis of genome-wide association scans from 14 studies totalling 28,141 participants of European descent, resulting in identification of 954 SNPs distributed across nine loci that exceeded the threshold of genome-wide significance, five of which are novel. Overall, the common variants associated with serum uric acid levels fall in the following nine regions: SLC2A9 (p=5.2×10⁻²⁰¹), ABCG2 (p=3.1×10⁻²⁶), SLC17A1 (p=3.0×10⁻¹⁴), SLC22A11 (p=6.7×10⁻¹⁴), SLC22A12 (p=2.0×10⁻⁹), SLC16A9 (p=1.1×10⁻⁸), GCKR (p=1.4×10⁻⁹), LRRC16A (p=8.5×10⁻⁹), and near PDZK1 (p=2.7×10⁻⁹). Identified variants were analyzed for gender differences. We found that the minor allele for rs734553 in SLC2A9 has greater influence in lowering uric acid levels in women and the minor allele of rs2231142 in ABCG2 elevates uric acid levels more strongly in men compared to women. To further characterize the identified variants, we analyzed their association with a panel of metabolites. rs12356193 within SLC16A9 was associated with DL-carnitine (p=4.0×10⁻²⁶) and propionyl-L-carnitine (p=5.0×10⁻⁸) concentrations, which in turn were associated with serum UA levels (p=1.4×10⁻⁵⁷ and p=8.1×10⁻⁵⁴, respectively), forming a triangle between SNP, metabolites, and UA levels. Taken together, these associations highlight additional pathways that are important in the regulation of serum uric acid levels and point toward novel potential targets for pharmacological intervention to prevent or treat hyperuricemia. In addition, these findings strongly support the hypothesis that transport proteins are key in regulating serum uric acid levels.     

Adaptation of the base-paired double-helix molecular architecture to extreme pressure

The behaviour of the d(GGTATACC) oligonucleotide has been investigated by X-ray crystallography at 295K in the range from ambient pressure to 2GPa (~20000atm). Four 3D-structures of the A-DNA form (at ambient pressure, 0.55, 1.09 and 1.39GPa) were refined at 1.60 or 1.65Å resolution. In addition to the diffraction pattern of the A-form, the broad meridional streaks previously explained by occluded B-DNA octamers within the channels of the crystalline A-form matrix were observed up to at least 2GPa. This work highlights an important property of nucleic acids, their capability to withstand very high pressures, while keeping in such conditions a nearly invariant geometry of base pairs that store and carry genetic information. The double-helix base-paired architecture behaves as a molecular spring, which makes it especially adapted to very harsh conditions. These features may have contributed to the emergence of a RNA World at prebiotic stage.     

Meta-Analysis of Genome-Wide Scans for Human Adult Stature Identifies Novel Loci and Associations with Measures of Skeletal Frame Size

Recent genome-wide (GW) scans have identified several independent loci affecting human stature, but their contribution through the different skeletal components of height is still poorly understood. We carried out a genome-wide scan in 12,611 participants, followed by replication in an additional 7,187 individuals, and identified 17 genomic regions with GW-significant association with height. Of these, two are entirely novel (rs11809207 in CATSPER4, combined P-value=6.1×10⁻⁸ and rs910316 in TMED10, P-value=1.4×10⁻⁷) and two had previously been described with weak statistical support (rs10472828 in NPR3, P-value=3×10⁻⁷ and rs849141 in JAZF1, P-value=3.2×10⁻¹¹). One locus (rs1182188 at GNA12) identifies the first height eQTL. We also assessed the contribution of height loci to the upper- (trunk) and lower-body (hip axis and femur) skeletal components of height. We find evidence for several loci associated with trunk length (including rs6570507 in GPR126, P-value=4×10⁻⁵ and rs6817306 in LCORL, P-value=4×10⁻⁴), hip axis length (including rs6830062 at LCORL, P-value=4.8×10⁻⁴ and rs4911494 at UQCC, P-value=1.9×10⁻⁴), and femur length (including rs710841 at PRKG2, P-value=2.4×10⁻⁵ and rs10946808 at HIST1H1D, P-value=6.4×10⁻⁶). Finally, we used conditional analyses to explore a possible differential contribution of the height loci to these different skeletal size measurements. In addition to validating four novel loci controlling adult stature, our study represents the first effort to assess the contribution of genetic loci to three skeletal components of height. Further statistical tests in larger numbers of individuals will be required to verify if the height loci affect height preferentially through these subcomponents of height.     

Isolation and Characterization of Temperate Bacteriophages of Clostridium difficile

The lack of information on bacteriophages of Clostridium difficile prompted this study. Three of 56 clinical C. difficile isolates yielded double-stranded DNA phages C2, C5, C6, and C8 upon induction. Superinfection and DNA analyses revealed relatedness between the phages, while partial sequencing of C2 showed nucleotide homology to the sequenced C. difficile strain CD630.