Functional Annotation of Genes Overlapping Copy Number Variants in Autistic Patients: Focus on Axon Pathfinding

We have used Gene Ontology (GO) and pathway analyses to uncover the common functions associated to the genes overlapping Copy Number Variants (CNVs) in autistic patients. Our source of data were four published studies [1-4]. We first applied a two-step enrichment strategy for autism-specific genes. We fished out from the four mentioned studies a list of 2928 genes overall overlapping 328 CNVs in patients and we first selected a sub-group of 2044 genes after excluding those ones that are also involved in CNVs reported in the Database of Genomic Variants (enrichment step 1). We then selected from the step 1-enriched list a sub-group of 514 genes each of which was found to be deleted or duplicated in at least two patients (enrichment step 2). The number of statistically significant processes and pathways identified by the Database for Annotation, Visualization and Integrated Discovery and Ingenuity Pathways Analysis softwares with the step 2-enriched list was significantly higher compared to the step 1-enriched list. In addition, statistically significant GO terms, biofunctions and pathways related to nervous system development and function were exclusively identified by the step 2-enriched list of genes. Interestingly, 21 genes were associated to axon growth and pathfinding. The latter genes and other ones associated to nervous system in this study represent a new set of autism candidate genes deserving further investigation. In summary, our results suggest that the autism’s “connectivity genes” in some patients affect very early phases of neurodevelopment, i.e., earlier than synaptogenesis

Myc Promoter-Binding Protein-1 (MBP-1) Is a Novel Potential Prognostic Marker in Invasive Ductal Breast Carcinoma

Background Alpha-enolase is a glycolytic enzyme that catalyses the formation of phosphoenolpyruvate in the cell cytoplasm. \u3b1-Enolase and the predominantly nuclear Myc promoter-binding protein-1 (MBP-1) originate from a single gene through the alternative use of translational starting sites. MBP-1 binds to the P2 c-myc promoter and competes with TATA-box binding protein (TBP) to suppress gene transcription. Although several studies have shown an antiproliferative effect of MBP-1 overexpression on several human cancer cells, to date detailed observations of \u3b1-enolase and MBP-1 relative expression in primary tumors versus normal tissues and their correlation with clinicopathological features have not been undertaken. Methodology and Findings We analyzed \u3b1-enolase and MBP-1 expression in normal breast epithelium and primary invasive ductal breast carcinoma (IDC) from 177 patients by Western blot and immunohistochemical analyses, using highly specific anti-\u3b1-enolase monoclonal antibodies. A significant increase in the expression of cytoplasmic \u3b1-enolase was observed in 98% of the tumors analysed, compared to normal tissues. Nuclear MBP-1 was found in almost all the normal tissues while its expression was retained in only 35% of the tumors. Statistically significant associations were observed among the nuclear expression of MBP-1 and ErbB2 status, Ki-67 expression, node status and tumor grade. Furthermore MBP-1 expression was associated with good survival of patients with IDC. Conclusions MBP-1 functions in repressing c-myc gene expression and the results presented indicate that the loss of nuclear MBP-1 expression in a large number of IDC may be a critical step in the development and progression of breast cancer and a predictor of adverse outcome. Nuclear MBP-1 appears to be a novel and valuable histochemical marker with potential prognostic value in breast cancer

Transgenic Mice for a Tamoxifen-Induced, Conditional Expression of the Cre Recombinase in Osteoclasts

Background: Studies on osteoclasts, the bone resorbing cells, have remained limited due to the lack of transgenic mice allowing the conditional knockout of genes in osteoclasts at any time during development or adulthood. Methodology/Principal Finding: We report here on the generation of transgenic mice which specifically express a tamoxifen-inducible Cre recombinase in osteoclasts. These mice, generated on C57BL/6 and FVB background, express a fusion Cre recombinase-ERT2 protein whose expression is driven by the promoter of cathepsin K (CtsK), a gene highly expressed in osteoclasts. We tested the cellular specificity of Cre activity in CtsKCreERT2 strains by breeding with Rosa26LacZ reporter mice. PCR and histological analyses of the CtsKCreERT2LacZ positive adult mice and E17.5 embryos show that Cre activity is restricted largely to bone tissue. In vitro, primary osteoclasts derived from the bone marrow of CtsKCreERT2+/2LacZ+/2 adult mice show a Cre-dependent b-galactosidase activity after tamoxifen stimulation

Development of standards of internal financial control of the enterprise

В статті аналізується стан законодавства, що регулює здійснення внутрішнього контролю на підприємствах. Обґрунтовується створення внутрішніх стандартів контролю для підвищення ефективності організації внутрішнього контролю на підприємствах.The article analyzes the state law governing the internal control in enterprises. Substantiated creation of internal control standards to improve the efficiency of internal control in enterprises

Tissue distribution of functional Cre recombinase in CtsKCreERT2LacZ E17.5 embryos.

<p>CtsKCreERT2LacZ and wild-type females were mated with ROSA26 males. Pregnant females were treated with tamoxifen as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037592#s3" target="_blank">materials and methods</a>. At day E17.5, the embryos were collected and analyzed for β-galactosidase activity (X-gal staining) as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037592#s3" target="_blank">materials and methods</a> using embryos with various genotypes.</p

Cre-mediated β-Galactosidase activity in osteoclasts derived from bone marrow of CtsKCreERT2/LacZ-positive transgenic mice.

<p>Primary osteoclasts were differentiated from bone marrow of tibias and femurs of CtsKCreERT2+/−LacZ+/− strain #4 as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037592#s3" target="_blank">materials and methods</a>. They were then treated with 1 µM 4-OHT or EtOH alone. After 24 hours, they were then treated to detect their β-galactosidase activity (A) or their TRAcP activity (B). The arrows indicate multinucleated osteoclasts.</p

Expression of Cre and β-galactosidase in adult mouse tissues.

<p>β-galactosidase and Cre expression in organs and bones of CtsKCreERT2+/−LacZ+/− (strain #4).Total RNAs were isolated from various tissues and expression was detected by semi quantitative (A) and quantitative (B) PCR as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037592#s3" target="_blank">materials and methods</a>. GAPDH was used as internal control and normalization.</p