Quality control and integration of genotypes from two calling pipelines for whole genome sequence data in the Alzheimer's disease sequencing project

The Alzheimer's Disease Sequencing Project (ADSP) performed whole genome sequencing (WGS) of 584 subjects from 111 multiplex families at three sequencing centers. Genotype calling of single nucleotide variants (SNVs) and insertion-deletion variants (indels) was performed centrally using GATK-HaplotypeCaller and Atlas V2. The ADSP Quality Control (QC) Working Group applied QC protocols to project-level variant call format files (VCFs) from each pipeline, and developed and implemented a novel protocol, termed “consensus calling,” to combine genotype calls from both pipelines into a single high-quality set. QC was applied to autosomal bi-allelic SNVs and indels, and included pipeline-recommended QC filters, variant-level QC, and sample-level QC. Low-quality variants or genotypes were excluded, and sample outliers were noted. Quality was assessed by examining Mendelian inconsistencies (MIs) among 67 parent-offspring pairs, and MIs were used to establish additional genotype-specific filters for GATK calls. After QC, 578 subjects remained. Pipeline-specific QC excluded ~12.0% of GATK and 14.5% of Atlas SNVs. Between pipelines, ~91% of SNV genotypes across all QCed variants were concordant; 4.23% and 4.56% of genotypes were exclusive to Atlas or GATK, respectively; the remaining ~0.01% of discordant genotypes were excluded. For indels, variant-level QC excluded ~36.8% of GATK and 35.3% of Atlas indels. Between pipelines, ~55.6% of indel genotypes were concordant; while 10.3% and 28.3% were exclusive to Atlas or GATK, respectively; and ~0.29% of discordant genotypes were. The final WGS consensus dataset contains 27,896,774 SNVs and 3,133,926 indels and is publicly available.     

Structure of the ovaries and follicular epithelium morphogenesis in Drosophila and its kin

In insects, the ovarian follicular epithelium morphogenesis has been intensively studied and best characterized in the fruit fly, Drosophila melanogaster. It is well established that initially identical somatic follicular cells (FCs) form a simple epithelium overlying the germline cells, but during oogenesis, they diversify into a number of morphologically distinct subpopulations each responsible for creating specific eggshell structures. In addition, some FC subpopulations (e.g. polar cells) are indispensable in establishing antero-posterior and dorso-ventral ovarian follicle axes and patterning of the developing embryo. The morphological and molecular changes that occur during follicular epithelium morphogenesis in Drosophila are frequently considered as a paradigm of the FC diversification in all flies. However, recent comparative studies indicate that, in dipterans, the functioning of the ovarian follicles is diverse, group-specific and may significantly differ from the Drosophila model system. We discuss the similarities and differences of the ovary structure and follicular epithelium morphogenesis in different dipteran groups and put them into a phylognetic context. We suggest that the migratory activity of the FCs represents an evolutionary novelty that evolved in the ancestors of higher dipterans (Brachycera). Subsequently, during evolution of this subgroup, the number of migrating FC subpopulations has gradually increased from one (in Orthorrhapha) to four (in Cyclorrhapha).     

Proteolytic Remodeling of the Synaptic Cell Adhesion Molecules (CAMs) by Metzincins in Synaptic Plasticity

Cell adhesion molecules participate in the formation, maturation, function and plasticity of synaptic connections. The growing body of evidence indicates that in the regulation of the synaptic plasticity, in which these molecules play pivotal role, also the proteolytic processes are involved. This review focuses on extracellular proteolysis of the cell adhesion molecules by specific subgroup of the matrix metalloproteinases, a disintegrin and metalloproteases and a disintegrin and metalloproteinase with thrombospondin motifs, jointly referred to as metzincins, in driving coordinated synaptic structural and functional modifications underlying synaptic plasticity in the adult brain.     

Anti-HmuY Antibodies Specifically Recognize Porphyromonas gingivalis HmuY Protein but Not Homologous Proteins in Other Periodontopathogens

Given the emerging evidence of an association between periodontal infections and systemic conditions, the search for specific methods to detect the presence of P. gingivalis, a principal etiologic agent in chronic periodontitis, is of high importance. The aim of this study was to characterize antibodies raised against purified P. gingivalis HmuY protein and selected epitopes of the HmuY molecule. Since other periodontopathogens produce homologs of HmuY, we also aimed to characterize responses of antibodies raised against the HmuY protein or its epitopes to the closest homologous proteins from Prevotella intermedia and Tannerella forsythia. Rabbits were immunized with purified HmuY protein or three synthetic, KLH-conjugated peptides, derived from the P. gingivalis HmuY protein. The reactivity of anti-HmuY antibodies with purified proteins or bacteria was determined using Western blotting and ELISA assay. First, we found homologs of P. gingivalis HmuY in P. intermedia (PinO and PinA proteins) and T. forsythia (Tfo protein) and identified corrected nucleotide and amino acid sequences of Tfo. All proteins were overexpressed in E. coli and purified using ion-exchange chromatography, hydrophobic chromatography and gel filtration. We demonstrated that antibodies raised against P. gingivalis HmuY are highly specific to purified HmuY protein and HmuY attached to P. gingivalis cells. No reactivity between P. intermedia and T. forsythia or between purified HmuY homologs from these bacteria and anti-HmuY antibodies was detected. The results obtained in this study demonstrate that P. gingivalis HmuY protein may serve as an antigen for specific determination of serum antibodies raised against this bacterium.     

Structure,function and DNA composition of Saccharomyces cerevisiae chromatin loops

Recent localization of cohesin association regions along the yeast chromatin fibre suggests that compositional variability of DNA in yeast is related to the function and organization of the chromosomal loops. The bases of the loops, where the chromatin fibre is attached to the chromosomal axis, are AT-rich, bind cohesin, and are flanked by genes transcribed convergently. The hotspots of meiotic recombination are mainly found in the GC-rich parts of the loops, ‘external’ with respect to the chromosomal axis, frequently in the vicinity of the promoters of divergently transcribed genes. There are two possible reasons why the regions of the hotspots of recombination were enriched in GC content during evolution. One is a biased repair of recombination intermediates, and the second is a selective advantage due to an increased chromatin accessibility, which may have the carriers of GC-enriched alleles over the carriers of AT-rich alleles.     

Cell cycle analysis of human peripheral blood T lymphocytes in long-term culture

We have studied the cell cycle of resting T lymphocytes from long-term (LT) cultures following stimulation with phytohemagglutinin (PHA) and recombinant Interleukin 2 (IL-2). We examined the kinetics of entry into S phase by autoradiography, the accumulation of cellular RNA by microfluorometric techniques, and ultrastructural morphology by electron microscopy. In addition, we examined the expression at the mRNA level of six cell cycle-dependent growth-regulated genes (c-fos, c-myc, KC-1, JE-3, vimentin, and histone H3). We show that T lymphocytes of LT cultures respond differently to mitogenic stimulation than the T lymphocytes of freshly isolated peripheral blood mononuclear cell cultures. At the ultrastructural, biochemical, and molecular levels, resting T lymphocytes of LT cultures can be distinguished from physiological (G0) lymphocytes of peripheral blood.     

Mechanism of microwave sterilization in the dry state

With an automated computerized temperature control and a specialized temperature measurement system, dry spores of Bacillus subtilis subsp. niger were treated with heat simultaneously in a convection dry-heat oven and a microwave oven. The temperature of the microwave oven was monitored such that the temperature profiles of the spore samples in both heat sources were nearly identical. Under these experimental conditions, we unequivocally demonstrated that the mechanism of sporicidal action of the microwaves was caused solely by thermal effects. Nonthermal effects were not significant in a dry microwave sterilization process. Both heating systems showed that a dwelling time of more than 45 min was required to sterilize 10(5) inoculated spores in dry glass vials at 137 degrees C. The D values of both heating systems were 88, 14, and 7 min at 117, 130, and 137 degrees C, respectively. The Z value was estimated to be 18 degrees C.