Pemphigus Vulgaris Autoantibody Profiling by Proteomic Technique

Pemphigus vulgaris (PV) is a mucocutaneous blistering disease characterized by IgG autoantibodies against the stratified squamous epithelium. Current understanding of PV pathophysiology does not explain the mechanism of acantholysis in patients lacking desmoglein antibodies, which justifies a search for novel targets of pemphigus autoimmunity. We tested 264 pemphigus and 138 normal control sera on the multiplexed protein array platform containing 701 human genes encompassing many known keratinocyte cell-surface molecules and members of protein families targeted by organ-non-specific PV antibodies. The top 10 antigens recognized by the majority of test patients’ sera were proteins encoded by the DSC1, DSC3, ATP2C1, PKP3, CHRM3, COL21A1, ANXA8L1, CD88 and CHRNE genes. The most common combinations of target antigens included at least one of the adhesion molecules DSC1, DSC3 or PKP3 and/or the acetylcholine receptor CHRM3 or CHRNE with or without the MHC class II antigen DRA. To identify the PV antibodies most specific to the disease process, we sorted the data based on the ratio of patient to control frequencies of antigen recognition. The frequency of antigen recognition by patients that exceeded that of control by 10 and more times were the molecules encoded by the CD33, GP1BA, CHRND, SLC36A4, CD1B, CD32, CDH8, CDH9, PMP22 and HLA-E genes as well as mitochondrial proteins encoded by the NDUFS1, CYB5B, SOD2, PDHA1 and FH genes. The highest specificity to PV showed combinations of autoantibodies to the calcium pump encoded by ATP2C1 with C5a receptor plus DSC1 or DSC3 or HLA-DRA. The results identified new targets of pemphigus autoimmunity. Novel autoantibody signatures may help explain individual variations in disease severity and treatment response, and serve as sensitive and specific biomarkers for new diagnostic assays in PV patients.     

Production of extracellular PETase from Ideonella sakaiensis using sec-dependent signal peptides in E. coli

Poly(ethylene terephthalate) (PET) is the most commonly used polyester polymer resin in fabrics and storage materials, and its accumulation in the environment is a global problem. The ability of PET hydrolase from Ideonella sakaiensis 201-F6 (IsPETase) to degrade PET at moderate temperatures has been studied extensively. However, due to its low structural stability and solubility, it is difficult to apply standard laboratory-level IsPETase expression and purification procedures in industry. To overcome this difficulty, the expression of IsPETase can be improved by using a secretion system. This is the first report on the production of an extracellular IsPETase, active against PET film, using Sec-dependent translocation signal peptides from E. coli. In this work, we tested the effects of fusions of the Sec-dependent and SRP-dependent signal peptides from E. coli secretory proteins into IsPETase, and successfully produced the extracellular enzyme using pET22b-SP_MalE:IsPETase and pET22b-SP_LamB:IsPETase expression systems. We also confirmed that the secreted IsPETase has PET-degradation activity. The work will be used for development of a new E. coli strain capable of degrading and assimilating PET in its culture medium.     

Correlation analysis for the incubation period of prion disease

Previous studies have shown that genetic quantitative trait loci (QTL), strain barriers, inoculation dose and inoculation method modulate the incubation period of prion diseases. We examined the relationship between a diverse set of physical, genetic and immunological characteristics and the incubation period of prion disease using correlation analyses. We found that incubation period was highly correlated with brain weight. In addition, mean corpuscular volume and cell size were strongly correlated with incubation period, indicating that the physical magnitude of prion-infected organs or individual cells may be important in determining the incubation period. Given the same prion inoculation dose, animals with a lower brain weight, mean corpuscular volume or cell size may experience more virulent disease, as the effective concentration of abnormal prion, which might regulate the attachment rate of prions to aggregates, is increased with smaller capacity of brains and cells. This is partly consistent with previous theoretical modeling. The strong correlations between incubation period and physical properties of the brain and cells in this study suggest that the mechanism underlying prion disease pathology may be physical, indicating that the incubation process is governed by simple chemical stoichiometry.     

Solidification/Stabilization of Cresol-Contaminated Soil: Mechanical and Leaching Behavior

In this research, the effects of a solidification/stabilization (S/S) technique on the remediation of cresol-contaminated soil were investigated. The soil samples were collected from the Tehran Oil Refinery (TOR) in Ray District, Iran. Cresols are hazardous chemicals whose exposure at high dosage results in irreparable damage to animal and human health. S/S process progresses through physicochemical reactions, reducing the leachability of a waste as well as changing its physical properties in order to encapsulate the contaminants and form a solid material. The samples were spiked by certain concentrations of cresols including meta-, ortho-, and para-isomers. The treatment process was conducted using Portland cement as the binder and modified bentonite as a stabilizer. The efficiency of the S/S technique was assessed by the unconfined compressive strength (UCS) test and toxicity characteristic leaching procedure (TCLP) test. The results of the tests showed that adding 20% and 30% Portland cement in combination with 15% and 30% modified bentonite to contaminated samples increased the 28-day compressive strength of S/S blocks to a range of 2.44 to 3.08 MPa. In fact, increasing the proportion of cement in the mix design resulted in enhanced compressive strength, while modified bentonite inversely affected samples’ strength. Regarding leaching behavior, adding organophilic clay to polluted samples noticeably declined cresol's concentrations in leachate. Overall efficiency of S/S in terms of leaching was about 70%, which was obtained by adding 20% cement and 30% modified clay. A logarithmic relationship between leaching percent and modified clay ratio was also detected with high logarithmic and linear correlation coefficients of 0.96 and 0.9, respectively, hence presenting the efficiency of S/S in stabilizing the cresols in the samples.     

Gene-Level,but Not Chromosome-Wide,Divergence between a Very Young House Fly Proto-Y Chromosome and Its Homologous Proto-X Chromosome

X and Y chromosomes are usually derived from a pair of homologous autosomes, which then diverge from each other over time. Although Y-specific features have been characterized in sex chromosomes of various ages, the earliest stages of Y chromosome evolution remain elusive. In particular, we do not know whether early stages of Y chromosome evolution consist of changes to individual genes or happen via chromosome-scale divergence from the X. To address this question, we quantified divergence between young proto-X and proto-Y chromosomes in the house fly, Musca domestica. We compared proto-sex chromosome sequence and gene expression between genotypic (XY) and sex-reversed (XX) males. We find evidence for sequence divergence between genes on the proto-X and proto-Y, including five genes with mitochondrial functions. There is also an excess of genes with divergent expression between the proto-X and proto-Y, but the number of genes is small. This suggests that individual proto-Y genes, but not the entire proto-Y chromosome, have diverged from the proto-X. We identified one gene, encoding an axonemal dynein assembly factor (which functions in sperm motility), that has higher expression in XY males than XX males because of a disproportionate contribution of the proto-Y allele to gene expression. The upregulation of the proto-Y allele may be favored in males because of this gene’s function in spermatogenesis. The evolutionary divergence between proto-X and proto-Y copies of this gene, as well as the mitochondrial genes, is consistent with selection in males affecting the evolution of individual genes during early Y chromosome evolution.     

Functional analyses of two syntaxin-like SNARE genes,GzSYN1 and GzSYN2, in the ascomycete Gibberella zeae

We identified two syntaxin-like SNARE genes, named GzSYN1 and GzSYN2, from the plant pathogenic ascomycete Gibberella zeae, and characterized the functions and cellular localization of these genes. The GzSYN1 deletion mutant (Δgzsyn1) had 71% reduced hyphal growth compared to the wild-type strain, but produced perithecia with normal ascospores. Δgzsyn2 had the same hyphal growth rate as the wild-type, but completely lost both self and female fertility. When Δgzsyn2 was spermatized for Δmat1-1 or Δmat1-2 strains, it retained its male fertility, but the ascus shape was abnormal and ascospore delimitation was delayed. The Δgzsyn1 and Δgzsyn2 virulence on barley was reduced by 67% and 75%, respectively, compared to the wild-type. The GFP::GzSYN1 fusion protein was localized in vesicles, vacuoles, plasma membranes, and septa, whereas GFP::GzSYN2 was found only in plasma membranes and septa. These results suggest that syntaxins have key roles in fungal development and virulence in G. zeae.