Immunoproteomic analysis of bacterial proteins of Actinobacillus pleuropneumoniae serotype 1

Background

Actinobacillus pleuropneumoniae (APP) is one of the most important swine pathogens worldwide. Identification and characterization of novel antigenic APP vaccine candidates are underway. In the present study, we use an immunoproteomic approach to identify APP protein antigens that may elicit an immune response in serotype 1 naturally infected swine and serotype 1 virulent strain S259-immunized rabbits.

Results

Proteins from total cell lysates of serotype 1 APP were separated by two-dimensional electrophoresis (2DE). Western blot analysis revealed 21 immunoreactive protein spots separated in the pH 4-7 range and 4 spots in the pH 7-11 range with the convalescent sera from swine; we found 5 immunoreactive protein spots that separated in the pH 4-7 range and 2 in the pH 7-11 range with hyperimmune sera from S259-immunized rabbits. The proteins included the known antigens ApxIIA, protective surface antigen D15, outer membrane proteins P5, subunit NqrA. The remaining antigens are being reported as immunoreactive proteins in APP for the first time, to our knowledge.

Conclusions

We identified a total of 42 immunoreactive proteins of the APP serotype 1 virulent strain S259 which represented 32 different proteins, including some novel immunoreactive factors which could be researched as vaccine candidates.     

Eight common genetic variants associated with serum DHEAS levels suggest a key role in ageing mechanisms

Dehydroepiandrosterone sulphate (DHEAS) is the most abundant circulating steroid secreted by adrenal glands--yet its function is unknown. Its serum concentration declines significantly with increasing age, which has led to speculation that a relative DHEAS deficiency may contribute to the development of common age-related diseases or diminished longevity. We conducted a meta-analysis of genome-wide association data with 14,846 individuals and identified eight independent common SNPs associated with serum DHEAS concentrations. Genes at or near the identified loci include ZKSCAN5 (rs11761528; p = 3.15 × 10(-36)), SULT2A1 (rs2637125; p = 2.61 × 10(-19)), ARPC1A (rs740160; p = 1.56 × 10(-16)), TRIM4 (rs17277546; p = 4.50 × 10(-11)), BMF (rs7181230; p = 5.44 × 10(-11)), HHEX (rs2497306; p = 4.64 × 10(-9)), BCL2L11 (rs6738028; p = 1.72 × 10(-8)), and CYP2C9 (rs2185570; p = 2.29 × 10(-8)). These genes are associated with type 2 diabetes, lymphoma, actin filament assembly, drug and xenobiotic metabolism, and zinc finger proteins. Several SNPs were associated with changes in gene expression levels, and the related genes are connected to biological pathways linking DHEAS with ageing. This study provides much needed insight into the function of DHEAS.     

Proteome profile of the pipping muscle in broiler embryos

This study is the first proteomics analysis of the muscularis complexus (pipping muscle) in chicken (Gallus gallus) broiler embryos. We used differential detergent fractionation and nano-HPLC-MS/MS analysis to identify 676 proteins from all cellular components. The identified proteins were functionally classified in accordance with their involvement in various cellular activities.     

Mitochondrial C150T polymorphism increases the risk of cervical cancer and HPV infection

During a survey of control region (D-loop) sequence variances in 142 cervical cancer (CC) patients and 136 controls, all Chinese women, including both HPV-positive (human papillomavirus) and HPV-negative subjects, we determined that the C150T polymorphism increased the CC risk in a case-control study (OR=3.0, 95% CI=1.8-5.0, P<0.05). HPV-positive individuals were more likely to carry the C150T polymorphism than HPV-negative controls (OR=5.8, 95% CI=2.6-13.2, P=2.3×10(-5)). HPV-positive CC patients were more likely to carry the C150T polymorphism than HPV-negative controls (OR=4.9, 95% CI=2.6-9.3, P=9.9×10(-7)). In all subjects, an increased risk of HPV infection was also associated with the C150T polymorphism (OR=4.5, 95% CI=2.5-8.1, P=6.6×10(-7)). However, no significant difference in the frequency of other alleles was found at the variable sites in D146, D152, D310 and D514. These results indicated that the C150T polymorphism increased the risk of HPV infection and CC progression. Additionally, we assessed the association of mtDNA copy number with CC risk or the C150T polymorphism in 45 CC patients and 43 controls. There was no significant association of mtDNA copy number with CC risk or the C150T polymorphism. To the best of our knowledge, this is the first report to suggest that mtDNA C150T polymorphism was positively associated with HPV infection and subsequent CC risk among Chinese women.     

Conformational changes and bioactivity of lysozyme on binding to and desorption from magnetite nanoparticles

A fundamental understanding of the conformational behaviors of lysozyme during the process of adsorption and desorption has been studied using spectrophotometric techniques, and interpreted in terms of the secondary structures in this work. FTIR data show an increase in α-helix and β-sheet content when lysozyme interaction with magnetite nanoparticles (Fe(3)O(4) (PEG+CM-CTS) NPs) which indicates that the lysozyme would adopt a more compact conformation state. The mechanism of fluorescence quenching of lysozyme by magnetite nanoparticles is due to the formation of lysozyme-nanoparticles complex. High desorption of lysozyme from Fe(3)O(4) (PEG+CM-CTS) NPs were achieved using phosphate buffer solution (PBS) (20 mM, pH 5.0, 0.2 M NaCl), PBS (20 mM, pH 5.0, 0.5 M NaCl) and acetic acid (0.2 M, pH 4.0) as eluents. The alterations of lysozyme secondary structure on desorption from nanoparticles were confirmed by circular dichroism and fluorescence spectroscopy. Lysozymes desorbed by PBS (20mM, pH 5.0, 0.2M NaCl) and PBS (20mM, pH 5.0, 0.5M NaCl) retain high fraction of its native structure with negligible effect on its activity, and about 92.4% and 89.5% activity were retained upon desorption from nanoparticles, however, lysozyme desorbed by acetic acid (0.2 M, pH 4.0) solution showed significant conformational changes. The stability of NPs-conjugated protein and retention of higher activity may find useful applications in biotechnology ranging from enzyme immobilization to protein purification.     

An unexpected function of the Prader-Willi syndrome imprinting center in maternal imprinting in mice

Genomic imprinting is a phenomenon that some genes are expressed differentially according to the parent of origin. Prader-Willi syndrome (PWS) and Angelman syndrome (AS) are neurobehavioral disorders caused by deficiency of imprinted gene expression from paternal and maternal chromosome 15q11-q13, respectively. Imprinted genes at the PWS/AS domain are regulated through a bipartite imprinting center, the PWS-IC and AS-IC. The PWS-IC activates paternal-specific gene expression and is responsible for the paternal imprint, whereas the AS-IC functions in the maternal imprint by allele-specific repression of the PWS-IC to prevent the paternal imprinting program. Although mouse chromosome 7C has a conserved PWS/AS imprinted domain, the mouse equivalent of the human AS-IC element has not yet been identified. Here, we suggest another dimension that the PWS-IC also functions in maternal imprinting by negatively regulating the paternally expressed imprinted genes in mice, in contrast to its known function as a positive regulator for paternal-specific gene expression. Using a mouse model carrying a 4.8-kb deletion at the PWS-IC, we demonstrated that maternal transmission of the PWS-IC deletion resulted in a maternal imprinting defect with activation of the paternally expressed imprinted genes and decreased expression of the maternally expressed imprinted gene on the maternal chromosome, accompanied by alteration of the maternal epigenotype toward a paternal state spread over the PWS/AS domain. The functional significance of this acquired paternal pattern of gene expression was demonstrated by the ability to complement PWS phenotypes by maternal inheritance of the PWS-IC deletion, which is in stark contrast to paternal inheritance of the PWS-IC deletion that resulted in the PWS phenotypes. Importantly, low levels of expression of the paternally expressed imprinted genes are sufficient to rescue postnatal lethality and growth retardation in two PWS mouse models. These findings open the opportunity for a novel approach to the treatment of PWS.