Chicken genome analysis reveals novel genes encoding biotin-binding proteins related to avidin family

BACKGROUND: A chicken egg contains several biotin-binding proteins (BBPs), whose complete DNA and amino acid sequences are not known. In order to identify and characterise these genes and proteins we studied chicken cDNAs and genes available in the NCBI database and chicken genome database using the reported N-terminal amino acid sequences of chicken egg-yolk BBPs as search strings. RESULTS: Two separate hits showing significant homology for these N-terminal sequences were discovered. For one of these hits, the chromosomal location in the immediate proximity of the avidin gene family was found. Both of these hits encode proteins having high sequence similarity with avidin suggesting that chicken BBPs are paralogous to avidin family. In particular, almost all residues corresponding to biotin binding in avidin are conserved in these putative BBP proteins. One of the found DNA sequences, however, seems to encode a carboxy-terminal extension not present in avidin. CONCLUSION: We describe here the predicted properties of the putative BBP genes and proteins. Our present observations link BBP genes together with avidin gene family and shed more light on the genetic arrangement and variability of this family. In addition, comparative modelling revealed the potential structural elements important for the functional and structural properties of the putative BBP proteins

Attenuated expression of tenascin-c in ovalbumin-challenged STAT4-/- mice

<p>Abstract</p> <p>Background</p> <p>Asthma leads to structural changes in the airways, including the modification of extracellular matrix proteins such as tenascin-C. The role of tenascin-C is unclear, but it might act as an early initiator of airway wall remodelling, as its expression is increased in the mouse and human airways during allergic inflammation. In this study, we examined whether Th1 or Th2 cells are important regulators of tenascin-C in experimental allergic asthma utilizing mice with impaired Th1 (STAT4-/-) or Th2 (STAT6-/-) immunity.</p> <p>Methods</p> <p>Balb/c wildtype (WT), STAT4-/- and STAT6-/- mice were sensitized with intraperitoneally injected ovalbumin (OVA) followed by OVA or PBS airway challenge. Airway hyperreactivity (AHR) was measured and samples were collected. Real time PCR and immunohistochemistry were used to study cytokines and differences in the expression of tenascin-C. Tenascin-C expression was measured in human fibroblasts after treatment with TNF-α and IFN-γ <it>in vitro</it>.</p> <p>Results</p> <p>OVA-challenged WT mice showed allergic inflammation and AHR in the airways along with increased expression of TNF-α, IFN-γ, IL-4 and tenascin-C in the lungs. OVA-challenged STAT4-/- mice exhibited elevated AHR and pulmonary eosinophilia. The mRNA expression of TNF-α and IFN-γ was low, but the expression of IL-4 was significantly elevated in these mice. OVA-challenged STAT6-/- mice had neither AHR nor pulmonary eosinophilia, but had increased expression of mRNA for TNF-α, IFN-γ and IL-4. The expression of tenascin-C in the lungs of OVA-challenged STAT4-/- mice was weaker than in those of OVA-challenged WT and STAT6-/- mice suggesting that TNF-α and IFN-γ may regulate tenascin-C expression <it>in vivo</it>. The stimulation of human fibroblasts with TNF-α and IFN-γ induced the expression of tenascin-C confirming our <it>in vivo </it>findings.</p> <p>Conclusions</p> <p>Expression of tenascin-C is significantly attenuated in the airways of STAT4-/- mice, which may be due to the impaired secretion of TNF-α and IFN-γ in these mice.</p

Meta-analysis of genome-wide linkage studies of asthma and related traits

<p>Abstract</p> <p>Background</p> <p>Asthma and allergy are complex multifactorial disorders, with both genetic and environmental components determining disease expression. The use of molecular genetics holds great promise for the identification of novel drug targets for the treatment of asthma and allergy. Genome-wide linkage studies have identified a number of potential disease susceptibility loci but replication remains inconsistent. The aim of the current study was to complete a meta-analysis of data from genome-wide linkage studies of asthma and related phenotypes and provide inferences about the consistency of results and to identify novel regions for future gene discovery.</p> <p>Methods</p> <p>The rank based genome-scan meta-analysis (GSMA) method was used to combine linkage data for asthma and related traits; bronchial hyper-responsiveness (BHR), allergen positive skin prick test (SPT) and total serum Immunoglobulin E (IgE) from nine Caucasian asthma populations.</p> <p>Results</p> <p>Significant evidence for susceptibility loci was identified for quantitative traits including; BHR (989 pedigrees, n = 4,294) 2p12-q22.1, 6p22.3-p21.1 and 11q24.1-qter, allergen SPT (1,093 pedigrees, n = 4,746) 3p22.1-q22.1, 17p12-q24.3 and total IgE (729 pedigrees, n = 3,224) 5q11.2-q14.3 and 6pter-p22.3. Analysis of the asthma phenotype (1,267 pedigrees, n = 5,832) did not identify any region showing genome-wide significance.</p> <p>Conclusion</p> <p>This study represents the first linkage meta-analysis to determine the relative contribution of chromosomal regions to the risk of developing asthma and atopy. Several significant results were obtained for quantitative traits but not for asthma confirming the increased phenotype and genetic heterogeneity in asthma. These analyses support the contribution of regions that contain previously identified asthma susceptibility genes and provide the first evidence for susceptibility loci on 5q11.2-q14.3 and 11q24.1-qter.</p

Development of a Tetrameric Streptavidin Mutein with Reversible Biotin Binding Capability: Engineering a Mobile Loop as an Exit Door for Biotin

A novel form of tetrameric streptavidin has been engineered to have reversible biotin binding capability. In wild-type streptavidin, loop3–4 functions as a lid for the entry and exit of biotin. When biotin is bound, interactions between biotin and key residues in loop3–4 keep this lid in the closed state. In the engineered mutein, a second biotin exit door is created by changing the amino acid sequence of loop7–8. This door is mobile even in the presence of the bound biotin and can facilitate the release of biotin from the mutein. Since loop7–8 is involved in subunit interactions, alteration of this loop in the engineered mutein results in an 11° rotation between the two dimers in reference to wild-type streptavidin. The tetrameric state of the engineered mutein is stabilized by a H127C mutation, which leads to the formation of inter-subunit disulfide bonds. The biotin binding kinetic parameters (koff of 4.28×10−4 s−1 and Kd of 1.9×10−8 M) make this engineered mutein a superb affinity agent for the purification of biotinylated biomolecules. Affinity matrices can be regenerated using gentle procedures, and regenerated matrices can be reused at least ten times without any observable reduction in binding capacity. With the combination of both the engineered mutein and wild-type streptavidin, biotinylated biomolecules can easily be affinity purified to high purity and immobilized to desirable platforms without any leakage concerns. Other potential biotechnological applications, such as development of an automated high-throughput protein purification system, are feasible

Correlation of exhaled breath temperature with bronchial blood flow in asthma

In asthma elevated rates of exhaled breath temperature changes (Δe°T) and bronchial blood flow (Q(aw)) may be due to increased vascularity of the airway mucosa as a result of inflammation. We investigated the relationship of Δe°T with Q(aw )and airway inflammation as assessed by exhaled nitric oxide (NO). We also studied the anti-inflammatory and vasoactive effects of inhaled corticosteroid and β(2)-agonist. Δe°T was confirmed to be elevated (7.27 ± 0.6 Δ°C/s) in 19 asthmatic subjects (mean age ± SEM, 40 ± 6 yr; 6 male, FEV(1 )74 ± 6 % predicted) compared to 16 normal volunteers (4.23 ± 0.41 Δ°C/s, p < 0.01) (30 ± 2 yr) and was significantly increased after salbutamol inhalation in normal subjects (7.8 ± 0.6 Δ°C/ s, p < 0.05) but not in asthmatic patients. Q(aw), measured using an acetylene dilution method was also elevated in patients with asthma compared to normal subjects (49.47 ± 2.06 and 31.56 ± 1.6 μl/ml/min p < 0.01) and correlated with exhaled NO (r = 0.57, p < 0.05) and Δe°T (r = 0.525, p < 0.05). In asthma patients, Q(aw )was reduced 30 minutes after the inhalation of budesonide 400 μg (21.0 ± 2.3 μl/ml/min, p < 0.05) but was not affected by salbutamol. Δe°T correlates with Q(aw )and exhaled NO in asthmatic patients and therefore may reflect airway inflammation, as confirmed by the rapid response to steroids

The "Snacking Child" and its social network: some insights from an italian survey

<p>Abstract</p> <p>Background</p> <p>The hypothesis underlying this work is that the social network of a child might have an impact on the alimentary behaviors, in particular for what concerns snack consumption patterns.</p> <p>Methods</p> <p>1215 Italian children 6-10 ys old were interviewed using a CATI facility in January 2010. 608 "snackers" and 607 "no-snackers" were identified. Information regarding family composition, child and relatives BMI, mother perception of child weight, child, father and mother physical activity, TV watching, social network, leisure time habits and dietary habits of peers, were collected. Association of variables with the status of snacker was investigated using a multivariable logistic regression model.</p> <p>Results</p> <p>Snackers children seem to be part of more numerous social network (1.40 friends vs 1.14, p = 0.042) where the majority of peers are also eating snacks, this percentage being significantly higher (89.5 vs 76.3, p < 0.001) than in the "no-snacker" group. The snacking group is identified by the fact that it tends to practice at least 4 hours per week of physical activity (OR: 1.36, CI: 1.03-1.9). No evidence of an association between snacking consumption and overweight status has been shown by our study.</p> <p>Conclusions</p> <p>The snacking child has more active peer-to-peer social relationships, mostly related with sport activities. However, spending leisure time in sportive activities implies being part of a social environment which is definitely a positive one from the point of view of obesity control, and indeed, no increase of overweight/obesity is seen in relation to snack consumption.</p

Hundreds of variants clustered in genomic loci and biological pathways affect human height

Most common human traits and diseases have a polygenic pattern of inheritance: DNA sequence variants at many genetic loci influence the phenotype. Genome-wide association (GWA) studies have identified more than 600 variants associated with human traits, but these typically explain small fractions of phenotypic variation, raising questions about the use of further studies. Here, using 183,727 individuals, we show that hundreds of genetic variants, in at least 180 loci, influence adult height, a highly heritable and classic polygenic trait. The large number of loci reveals patterns with important implications for genetic studies of common human diseases and traits. First, the 180 loci are not random, but instead are enriched for genes that are connected in biological pathways (P = 0.016) and that underlie skeletal growth defects (P < 0.001). Second, the likely causal gene is often located near the most strongly associated variant: in 13 of 21 loci containing a known skeletal growth gene, that gene was closest to the associated variant. Third, at least 19 loci have multiple independently associated variants, suggesting that allelic heterogeneity is a frequent feature of polygenic traits, that comprehensive explorations of already-discovered loci should discover additional variants and that an appreciable fraction of associated loci may have been identified. Fourth, associated variants are enriched for likely functional effects on genes, being over-represented among variants that alter amino-acid structure of proteins and expression levels of nearby genes. Our data explain approximately 10% of the phenotypic variation in height, and we estimate that unidentified common variants of similar effect sizes would increase this figure to approximately 16% of phenotypic variation (approximately 20% of heritable variation). Although additional approaches are needed to dissect the genetic architecture of polygenic human traits fully, our findings indicate that GWA studies can identify large numbers of loci that implicate biologically relevant genes and pathways.

Mast Cells Express 11 beta-hydroxysteroid Dehydrogenase Type 1: A Role in Restraining Mast Cell Degranulation:a role in restraining mast cell degranulation

Mast cells are key initiators of allergic, anaphylactic and inflammatory reactions, producing mediators that affect vascular permeability, angiogenesis and fibrosis. Glucocorticoid pharmacotherapy reduces mast cell number, maturation and activation but effects at physiological levels are unknown. Within cells, glucocorticoid concentration is modulated by the 11β-hydroxysteroid dehydrogenases (11β-HSDs). Here we show expression and activity of 11β-HSD1, but not 11β-HSD2, in mouse mast cells with 11β-HSD activity only in the keto-reductase direction, regenerating active glucocorticoids (cortisol, corticosterone) from inert substrates (cortisone, 11-dehydrocorticosterone). Mast cells from 11β-HSD1-deficient mice show ultrastructural evidence of increased activation, including piecemeal degranulation and have a reduced threshold for IgG immune complex-induced mast cell degranulation. Consistent with reduced intracellular glucocorticoid action in mast cells, levels of carboxypeptidase A3 mRNA, a glucocorticoid-inducible mast cell-specific transcript, are lower in peritoneal cells from 11β-HSD1-deficient than control mice. These findings suggest that 11β-HSD1-generated glucocorticoids may tonically restrain mast cell degranulation, potentially influencing allergic, anaphylactic and inflammatory responses

Prevention of recurrent sickness absence among employees with common mental disorders: design of a cluster-randomised controlled trial with cost-benefit and effectiveness evaluation

BACKGROUND: Common mental disorders, such as depression, anxiety disorder, and adjustment disorder, have emerged as a major public and occupational health problem in many countries. These disorders can have severe consequences such as absenteeism and work disability. Different interventions have been developed to improve the return-to-work of employees with common mental disorders, but still a large proportion of employees experiences health and work problems after their return-to-work. For this reason, the SHARP-at work intervention is developed to prevent a relapse of sickness absence among employees who have returned to work after a period of sickness absence because of common mental disorders. We aim to evaluate the effectiveness, cost-benefit and process of the intervention compared to care as usual. METHODS/DESIGN: The study is designed as a cluster-randomised controlled trial with randomisation at the level of the occupational physician. Employees who have returned to work after a period of sickness absence because of a common mental disorder are included in the study. Employees in the intervention group will receive the SHARP-at work intervention. The intervention focusses on active guidance of employees by occupational physicians during the first weeks of work after sickness absence. Employees in the control group will receive care as usual. Outcomes will be assessed at baseline and at 3, 6, and 12 months follow-up. The primary outcome is cumulative recurrent sickness absence days. Secondary outcome measures are mental health, work functioning, and coping. Adherence to the protocol, communication between stakeholders, and satisfaction with the treatment are the process measures assessed in both study groups. Cost-benefit is calculated from a societal perspective. Finally, prognostic factors for a relapse of sickness absence are investigated. DISCUSSION: This study goes beyond return-to-work by focussing on the prevention of recurrent sickness absence. The study incorporates not only outcomes on sickness absence and mental health but also on health-related work functioning. The results of this study can contribute to a further development of practice guidelines and the promotion of sustainable work participation. TRIAL REGISTRATION: NTR1963

Construction of Chimeric Dual-Chain Avidin by Tandem Fusion of the Related Avidins

BACKGROUND: Avidin is a chicken egg-white protein with high affinity to vitamin H, also known as D-biotin. Many applications in life science research are based on this strong interaction. Avidin is a homotetrameric protein, which promotes its modification to symmetrical entities. Dual-chain avidin, a genetically engineered avidin form, has two circularly permuted chicken avidin monomers that are tandem-fused into one polypeptide chain. This form of avidin enables independent modification of the two domains, including the two biotin-binding pockets; however, decreased yields in protein production, compared to wt avidin, and complicated genetic manipulation of two highly similar DNA sequences in the tandem gene have limited the use of dual-chain avidin in biotechnological applications. PRINCIPAL FINDINGS: To overcome challenges associated with the original dual-chain avidin, we developed chimeric dual-chain avidin, which is a tandem fusion of avidin and avidin-related protein 4 (AVR4), another member of the chicken avidin gene family. We observed an increase in protein production and better thermal stability, compared with the original dual-chain avidin. Additionally, PCR amplification of the hybrid gene was more efficient, thus enabling more convenient and straightforward modification of the dual-chain avidin. When studied closer, the generated chimeric dual-chain avidin showed biphasic biotin dissociation. SIGNIFICANCE: The improved dual-chain avidin introduced here increases its potential for future applications. This molecule offers a valuable base for developing bi-functional avidin tools for bioseparation, carrier proteins, and nanoscale adapters. Additionally, this strategy could be helpful when generating hetero-oligomers from other oligomeric proteins with high structural similarity