Hormad1 mutation disrupts synaptonemal complex formation, recombination, and chromosome segregation in mammalian meiosis

Meiosis is unique to germ cells and essential for reproduction. During the first meiotic division, homologous chromosomes pair, recombine, and form chiasmata. The homologues connect via axial elements and numerous transverse filaments to form the synaptonemal complex. The synaptonemal complex is a critical component for chromosome pairing, segregation, and recombination. We previously identified a novel germ cell-specific HORMA domain encoding gene, Hormad1, a member of the synaptonemal complex and a mammalian counterpart to the yeast meiotic HORMA domain protein Hop1. Hormad1 is essential for mammalian gametogenesis as knockout male and female mice are infertile. Hormad1 deficient (Hormad1-/-) testes exhibit meiotic arrest in the early pachytene stage, and synaptonemal complexes cannot be visualized by electron microscopy. Hormad1 deficiency does not affect localization of other synaptonemal complex proteins, SYCP2 and SYCP3, but disrupts homologous chromosome pairing. Double stranded break formation and early recombination events are disrupted in Hormad1-/- testes and ovaries as shown by the drastic decrease in the γH2AX, DMC1, RAD51, and RPA foci. HORMAD1 co-localizes with cH2AX to the sex body during pachytene. BRCA1, ATR, and γH2AX co-localize to the sex body and participate in meiotic sex chromosome inactivation and transcriptional silencing. Hormad1 deficiency abolishes γH2AX, ATR, and BRCA1 localization to the sex chromosomes and causes transcriptional de-repression on the X chromosome. Unlike testes, Hormad1-/- ovaries have seemingly normal ovarian folliculogenesis after puberty. However, embryos generated from Hormad1-/- oocytes are hyper- and hypodiploid at the 2 cell and 8 cell stage, and they arrest at the blastocyst stage. HORMAD1 is therefore a critical component of the synaptonemal complex that affects synapsis, recombination, and meiotic sex chromosome inactivation and transcriptional silencing. © 2010 Shin et al

iRFP is a real time marker for transformation based assays in high content screening

Anchorage independent growth is one of the hallmarks of oncogenic transformation. Here we show that infrared fluorescent protein (iRFP) based assays allow accurate and unbiased determination of colony formation and anchorage independent growth over time. This protocol is particularly compatible with high throughput systems, in contrast to traditional methods which are often labor-intensive, subjective to bias and do not allow further analysis using the same cells. Transformation in a single layer soft agar assay could be documented as early as 2 to 3 days in a 96 well format, which can be easily combined with standard transfection, infection and compound screening setups to allow for high throughput screening to identify therapeutic targets

Euler diagrams through the looking glass: From extent to intent

Extension and intension are two ways of indicating the fundamental meaning of a concept. The extent of a concept, C, is the set of objects which correspond to C whereas the intent of C is the collection of attributes that characterise it. Thus, intension denotes the set of objects corresponding to C without naming them individually. Mathematicians switch comfortably between these perspectives but the majority of logical diagrams deal exclusively in extension. Euler diagrams indicate sets using curves to depict their extent in a way that intuitively matches the relations between the sets. What happens when we use spatial diagrams to depict intension? What can we infer about the intension of a concept given its extension, and vice versa? We present the first steps towards addressing these questions by defining extensional and intensional Euler diagrams and translations between the two perspectives. We show that translation in either direction leads to a loss of information, yet preserves important semantic properties. To conclude, we explain how we expect further exploration of the relationship between the two perspectives could shed light on connections between diagrams, extension, intension, and well-matchedness

Measurement of the residual stress tensor in a compact tension weld specimen

Neutron diffraction measurements have been performed to determine the full residual stress tensor along the expected crack path in an austenitic stainless steel (Esshete 1250) compact tension weld specimen. A destructive slitting method was then implemented on the same specimen to measure the stress intensity factor profile associated with the residual stress field as a function of crack length. Finally deformations of the cut surfaces were measured to determine a contour map of the residual stresses in the specimen prior to the cut. The distributions of transverse residual stress measured by the three techniques are in close agreement. A peak tensile stress in excess of 600 MPa was found to be associated with an electron beam weld used to attach an extension piece to the test sample, which had been extracted from a pipe manual metal arc butt weld. The neutron diffraction measurements show that exceptionally high residual stress triaxiality is present at crack depths likely to be used for creep crack growth testing and where a peak stress intensity factor of 35 MPa√m was measured (crack depth of 21 mm). The neutron diffraction measurements identified maximum values of shear stress in the order of 50 MPa and showed that the principal stress directions were aligned to within ~20° of the specimen orthogonal axes. Furthermore it was confirmed that measurement of strains by neutron diffraction in just the three specimen orthogonal directions would have been sufficient to provide a reasonably accurate characterisation of the stress state in welded CT specimens

Novel Approaches to Inhibition of Gastric Acid Secretion

The gastric H,K-adenosine triphosphatase (ATPase) is the primary target for treatment of acid-related diseases. Proton pump inhibitors (PPIs) are weak bases composed of two moieties, a substituted pyridine with a primary pKa of about 4.0 that allows selective accumulation in the secretory canaliculus of the parietal cell, and a benzimidazole with a second pKa of about 1.0. Protonation of this benzimidazole activates these prodrugs, converting them to sulfenic acids and/or sulfenamides that react covalently with one or more cysteines accessible from the luminal surface of the ATPase. The maximal pharmacodynamic effect of PPIs as a group relies on cyclic adenosine monophosphate–driven H,K-ATPase translocation from the cytoplasm to the canalicular membrane of the parietal cell. At present, this effect can only be achieved with protein meal stimulation. Because of covalent binding, inhibitory effects last much longer than their plasma half-life. However, the short dwell-time of the drug in the blood and the requirement for acid activation impair their efficacy in acid suppression, particularly at night. All PPIs give excellent healing of peptic ulcer and produce good, but less than satisfactory, results in reflux esophagitis. PPIs combined with antibiotics eradicate Helicobacter pylori, but success has fallen to less than 80%. Longer dwell-time PPIs promise to improve acid suppression and hence clinical outcome. Potassium-competitive acid blockers (P-CABs) are another class of ATPase inhibitors, and at least one is in development. The P-CAB under development has a long duration of action even though its binding is not covalent. PPIs with a longer dwell time or P-CABs with long duration promise to address unmet clinical needs arising from an inability to inhibit nighttime acid secretion, with continued symptoms, delayed healing, and growth suppression of H. pylori reducing susceptibility to clarithromycin and amoxicillin. Thus, novel and more effective suppression of acid secretion would benefit those who suffer from acid-related morbidity, continuing esophageal damage and pain, nonsteroidal anti-inflammatory drug–induced ulcers, and nonresponders to H. pylori eradication

Evaluation of Errors Associated with Cutting-Induced Plasticity in Residual Stress Measurements Using the Contour Method

Cutting-induced plasticity can lead to elevated uncertainties in residual stress measurements made by the contour method. In this study plasticity-induced stress errors are numerically evaluated for a benchmark edge-welded beam to understand the underlying mechanism. Welding and cutting are sequentially simulated by finite element models which have been validated by previous experimental results. It is found that a cutting direction normal to the symmetry plane of the residual stress distribution can lead to a substantially asymmetrical back-calculated stress distribution, owing to cutting-induced plasticity. In general, the stresses at sample edges are most susceptible to error, particularly when the sample is restrained during cutting. Inadequate clamping (far from the plane of cut) can lead to highly concentrated plastic deformation in local regions, and consequently the back-calculated stresses have exceptionally high values and gradients at these locations. Furthermore, the overall stress distribution is skewed towards the end-of-cut side. Adequate clamping (close to the plane of cut) minimises errors in back-calculated stress which becomes insensitive to the cutting direction. For minimal constraint (i.e. solely preventing rigid body motion), the plastic deformation is relatively smoothly distributed, and an optimal cutting direction (i.e. cutting from the base material towards the weld region in a direction that falls within the residual stress symmetry plane) is identified by evaluating the magnitude of stress errors. These findings suggest that cutting process information is important for the evaluation of potential plasticity-induced errors in contour method results, and that the cutting direction and clamping strategy can be optimised with an understanding of their effects on plasticity and hence the back-calculated stresses

Urban energy consumption and CO2 emissions in Beijing: current and future

This paper calculates the energy consumption and CO2 emissions of Beijing over 2005–2011 in light of the Beijing’s energy balance table and the carbon emission coefficients of IPCC. Furthermore, based on a series of energy conservation planning program issued in Beijing, the Long-range Energy Alternatives Planning System (LEAP)-BJ model is developed to study the energy consumption and CO2 emissions of Beijing’s six end-use sectors and the energy conversion sector over 2012–2030 under the BAU scenario and POL scenario. Some results are found in this research: (1) During 2005–2011, the energy consumption kept increasing, while the total CO2 emissions fluctuated obviously in 2008 and 2011. The energy structure and the industrial structure have been optimized to a certain extent. (2) If the policies are completely implemented, the POL scenario is projected to save 21.36 and 35.37 % of the total energy consumption and CO2 emissions than the BAU scenario during 2012 and 2030. (3) The POL scenario presents a more optimized energy structure compared with the BAU scenario, with the decrease of coal consumption and the increase of natural gas consumption. (4) The commerce and service sector and the energy conversion sector will become the largest contributor to energy consumption and CO2 emissions, respectively. The transport sector and the industrial sector are the two most potential sectors in energy savings and carbon reduction. In terms of subscenarios, the energy conservation in transport (TEC) is the most effective one. (5) The macroparameters, such as the GDP growth rate and the industrial structure, have great influence on the urban energy consumption and carbon emissions

Transactivation of EGFR by LPS induces COX-2 expression in enterocytes

Necrotizing enterocolitis (NEC) is the leading cause of gastrointestinal morbidity and mortality in preterm infants. NEC is characterized by an exaggerated inflammatory response to bacterial flora leading to bowel necrosis. Bacterial lipopolysaccharide (LPS) mediates inflammation through TLR4 activation and is a key molecule in the pathogenesis of NEC. However, LPS also induces cyclooxygenase-2 (COX-2), which promotes intestinal barrier restitution through stimulation of intestinal cell survival, proliferation, and migration. Epidermal growth factor receptor (EGFR) activation prevents experimental NEC and may play a critical role in LPS-stimulated COX-2 production. We hypothesized that EGFR is required for LPS induction of COX-2 expression. Our data show that inhibiting EGFR kinase activity blocks LPS-induced COX-2 expression in small intestinal epithelial cells. LPS induction of COX-2 requires Src-family kinase signaling while LPS transactivation of EGFR requires matrix metalloprotease (MMP) activity. EGFR tyrosine kinase inhibitors block LPS stimulation of mitogen-activated protein kinase ERK, suggesting an important role of the MAPK/ERK pathway in EGFR-mediated COX-2 expression. LPS stimulates proliferation of IEC-6 cells, but this stimulation is inhibited with either the EGFR kinase inhibitor AG1478, or the selective COX-2 inhibitor Celecoxib. Taken together, these data show that EGFR plays an important role in LPS-induction of COX-2 expression in enterocytes, which may be one mechanism for EGF in inhibition of NEC

Effect of promoter architecture on the cell-to-cell variability in gene expression

According to recent experimental evidence, the architecture of a promoter, defined as the number, strength and regulatory role of the operators that control the promoter, plays a major role in determining the level of cell-to-cell variability in gene expression. These quantitative experiments call for a corresponding modeling effort that addresses the question of how changes in promoter architecture affect noise in gene expression in a systematic rather than case-by-case fashion. In this article, we make such a systematic investigation, based on a simple microscopic model of gene regulation that incorporates stochastic effects. In particular, we show how operator strength and operator multiplicity affect this variability. We examine different modes of transcription factor binding to complex promoters (cooperative, independent, simultaneous) and how each of these affects the level of variability in transcription product from cell-to-cell. We propose that direct comparison between in vivo single-cell experiments and theoretical predictions for the moments of the probability distribution of mRNA number per cell can discriminate between different kinetic models of gene regulation.Comment: 35 pages, 6 figures, Submitte