Rap1B promotes VEGF-induced endothelial permeability and is required for dynamic regulation of the endothelial barrier

Vascular endothelial growth factor (VEGF), a key angiogenic and permeability factor, plays an important role in new blood vessel formation. However, abnormal VEGF-induced VEGFR2 signaling leads to hyperpermeability. We have shown previously that Rap1, best known for promoting cell adhesion and vessel stability, is a critical regulator of VEGFR2-mediated angiogenic and shear-stress EC responses. To determine the role of Rap1 role in endothelial barrier dynamics, we examined vascular permeability in EC-specific Rap1A- and Rap1B-knockout mice, cell-cell junction remodeling and EC monolayer resistivity in Rap1-deficient ECs under basal, inflammatory or elevated VEGF conditions. Deletion of either Rap1 isoform impaired de novo adherens junction (AJ) formation and recovery from LPS-induced barrier disruption in vivo However, only Rap1A deficiency increased permeability in ECs and lung vessels. Interestingly, Rap1B deficiency attenuated VEGF-induced permeability in vivo and AJ remodeling in vitro Therefore, only Rap1A is required for the maintenance of normal vascular integrity. Importantly, Rap1B is the primary isoform essential for normal VEGF-induced EC barrier dissolution. Deletion of either Rap1 isoform protected against hyper permeability in the STZ-induced diabetes model, suggesting clinical implications for targeting Rap1 in pathologies with VEGF-induced hyperpermeability

Spectrum of JAG1 gene mutations in Polish patients with Alagille syndrome

Alagille syndrome (ALGS) is an autosomal dominant disorder characterized by developmental abnormalities in several organs including the liver, heart, eyes, vertebrae, kidneys, and face. The majority (90-94 %) of ALGS cases are caused by mutations in the JAG1 (JAGGED1) gene, and in a small percent of patients (∼1 %) mutations in the NOTCH2 gene have been described. Both genes are involved in the Notch signaling pathway. To date, over 440 different JAG1 gene mutations and ten NOTCH2 mutations have been identified in ALGS patients. The present study was conducted on a group of 35 Polish ALGS patients and revealed JAG1 gene mutations in 26 of them. Twenty-three different mutations were detected including 13 novel point mutations and six large deletions affecting the JAG1 gene. Review of all mutations identified to date in individuals from Poland allowed us to propose an effective diagnostic strategy based on the mutations identified in the reported patients of Polish descent. However, the distribution of mutations seen in this cohort was not substantively different than the mutation distribution in other reported populations

DNA damage and transcriptional regulation in iPSC-derived neurons from Ataxia Telangiectasia patients

Abstract Ataxia Telangiectasia (A-T) is neurodegenerative syndrome caused by inherited mutations inactivating the ATM kinase, a master regulator of the DNA damage response (DDR). What makes neurons vulnerable to ATM loss remains unclear. In this study we assessed on human iPSC-derived neurons whether the abnormal accumulation of DNA-Topoisomerase 1 adducts (Top1ccs) found in A-T impairs transcription elongation, thus favoring neurodegeneration. Furthermore, whether neuronal activity-induced immediate early genes (IEGs), a process involving the formation of DNA breaks, is affected by ATM deficiency. We found that Top1cc trapping by CPT induces an ATM-dependent DDR as well as an ATM-independent induction of IEGs and repression especially of long genes. As revealed by nascent RNA sequencing, transcriptional elongation and recovery were found to proceed with the same rate, irrespective of gene length and ATM status. Neuronal activity induced by glutamate receptors stimulation, or membrane depolarization with KCl, triggered a DDR and expression of IEGs, the latter independent of ATM. In unperturbed A-T neurons a set of genes (FN1, DCN, RASGRF1, FZD1, EOMES, SHH, NR2E1) implicated in the development, maintenance and physiology of central nervous system was specifically downregulated, underscoring their potential involvement in the neurodegenerative process in A-T patients

Prevalence of overweight and obesity in children aged 6–13 years—alarming increase in obesity in Cracow, Poland

This study in children aged 6–13 years (n = 1,499) was performed between October 2008 and March 2009. Height and weight measurements were taken to calculate BMI. The prevalence of overweight and obesity was determined by means of IOTF cut-offs with respect to age. Alarming is the fact that the percentage of obese children in Cracow increased dramatically from 1.04% in boys and 0.20% in girls in 1971 to 7% in boys and 3.6% in girls in 2009. In this report, a higher percentage of overweight boys was observed in rural boys (28.14%) than in urban ones (27.31%). Obesity was identified in an almost twice as high percentage of urban boys (7.78%) as in rural ones (3.52%). A higher percentage of overweight girls was registered in rural areas (16.49%) than in urban ones (16.09%). Obesity was prevailing in rural girls (4.12%) relative to their urban counterparts (3.44%). The highest number of overweight urban boys was diagnosed in the group of 12-year-olds (n = 48) and rural boys in the group of 10-year-olds (n = 39), as well as in urban girls aged 11 (n = 17) and rural girls aged 9 (n = 9). The highest number of obesity was observed in rural boys aged 12 (n = 3) and in urban boys aged 9 and 10 (n = 9 in both groups). In the group of girls, obesity prevailed in urban 9-year-olds (n = 5) and in rural 7-year-olds (n = 5). Conclusions: Overweight and obesity affect boys almost twice as frequently as girls. Obesity is twice as frequent in urban boys as in their rural peers

The novel gene Ny-1 on potato chromosome IX confers hypersensitive resistance to Potato virus Y and is an alternative to Ry genes in potato breeding for PVY resistance

Hypersensitive resistance (HR) is an efficient defense strategy in plants that restricts pathogen growth and can be activated during host as well as non-host interactions. HR involves programmed cell death and manifests itself in tissue collapse at the site of pathogen attack. A novel hypersensitivity gene, Ny-1, for resistance to Potato virus Y (PVY) was revealed in potato cultivar Rywal. This is the first gene that confers HR in potato plants both to common and necrotic strains of PVY. The locus Ny-1 mapped on the short arm of potato chromosome IX, where various resistance genes are clustered in Solanaceous genomes. Expression of HR was temperature-dependent in cv. Rywal. Strains PVYO and PVYN, including subgroups PVYNW and PVYNTN, were effectively localized when plants were grown at 20°C. At 28°C, plants were systemically infected but no symptoms were observed. In field trials, PVY was restricted to the inoculated leaves and PVY-free tubers were produced. Therefore, the gene Ny-1 can be useful for potato breeding as an alternative donor of PVY resistance, because it is efficacious in practice-like resistance conferred by Ry genes

The MRN complex is transcriptionally regulated by MYCN during neural cell proliferation to control replication stress

The MRE11/RAD50/NBS1 (MRN) complex is a major sensor of DNA double strand breaks, whose role in controlling faithful DNA replication and preventing replication stress is also emerging. Inactivation of the MRN complex invariably leads to developmental and/or degenerative neuronal defects, the pathogenesis of which still remains poorly understood. In particular, NBS1 gene mutations are associated with microcephaly and strongly impaired cerebellar development, both in humans and in the mouse model. These phenotypes strikingly overlap those induced by inactivation of MYCN, an essential promoter of the expansion of neuronal stem and progenitor cells, suggesting that MYCN and the MRN complex might be connected on a unique pathway essential for the safe expansion of neuronal cells. Here, we show that MYCN transcriptionally controls the expression of each component of the MRN complex. By genetic and pharmacological inhibition of the MRN complex in a MYCN overexpression model and in the more physiological context of the Hedgehog-dependent expansion of primary cerebellar granule progenitor cells, we also show that the MRN complex is required for MYCN-dependent proliferation. Indeed, its inhibition resulted in DNA damage, activation of a DNA damage response, and cell death in a MYCN- and replication-dependent manner. Our data indicate the MRN complex is essential to restrain MYCN-induced replication stress during neural cell proliferation and support the hypothesis that replication-born DNA damage is responsible for the neuronal defects associated with MRN dysfunctions.Cell Death and Differentiation advance online publication, 12 June 2015; doi:10.1038/cdd.2015.81

Binding Properties and Stability of the Ras-Association Domain of Rap1-GTP Interacting Adapter Molecule (RIAM)

The Rap1-GTP interacting adapter protein (RIAM) is an important protein in Rap1-mediated integrin activation. By binding to both Rap1 GTPase and talin, RIAM recruits talin to the cell membrane, thus facilitating talin-dependent integrin activation. In this article, we studied the role of the RIAM Ras-association (RA) and pleckstrin-homology (PH) domains in the interaction with Rap1. We found that the RA domain was sufficient for GTP-dependent interaction with Rap1B, and the addition of the PH domain did not change the binding affinity. We also detected GTP-independent interaction of Rap1B with the N-terminus of RIAM. In addition, we found that the PH domain stabilized the RA domain both in vitro and in cells

Myosin VIIA, Important for Human Auditory Function, Is Necessary for Drosophila Auditory Organ Development

BACKGROUND: Myosin VIIA (MyoVIIA) is an unconventional myosin necessary for vertebrate audition [1]-[5]. Human auditory transduction occurs in sensory hair cells with a staircase-like arrangement of apical protrusions called stereocilia. In these hair cells, MyoVIIA maintains stereocilia organization [6]. Severe mutations in the Drosophila MyoVIIA orthologue, crinkled (ck), are semi-lethal [7] and lead to deafness by disrupting antennal auditory organ (Johnston's Organ, JO) organization [8]. ck/MyoVIIA mutations result in apical detachment of auditory transduction units (scolopidia) from the cuticle that transmits antennal vibrations as mechanical stimuli to JO. PRINCIPAL FINDINGS: Using flies expressing GFP-tagged NompA, a protein required for auditory organ organization in Drosophila, we examined the role of ck/MyoVIIA in JO development and maintenance through confocal microscopy and extracellular electrophysiology. Here we show that ck/MyoVIIA is necessary early in the developing antenna for initial apical attachment of the scolopidia to the articulating joint. ck/MyoVIIA is also necessary to maintain scolopidial attachment throughout adulthood. Moreover, in the adult JO, ck/MyoVIIA genetically interacts with the non-muscle myosin II (through its regulatory light chain protein and the myosin binding subunit of myosin II phosphatase). Such genetic interactions have not previously been observed in scolopidia. These factors are therefore candidates for modulating MyoVIIA activity in vertebrates. CONCLUSIONS: Our findings indicate that MyoVIIA plays evolutionarily conserved roles in auditory organ development and maintenance in invertebrates and vertebrates, enhancing our understanding of auditory organ development and function, as well as providing significant clues for future research

Secular Trends in the Prevalence of Overweight and Obesity in Sicilian Schoolchildren Aged 11–13 Years During the Last Decade

The present study evaluates trends in the prevalence of overweight and obesity in relation to gender and area of residence between two cohorts of students aged 11–13 years in Sicily. The analysis was performed on 1,839 schoolchildren, with 924 and 915 children being studied in 1999–2001 and 2009–2010, respectively. The children who were enrolled during 2009–2010 had significantly higher body mass indexes (BMI), BMI z-scores, and waist circumferences than the children who were studied during 1999–2001 (p<0.0001 for all); these differences was also observed when the cohort was subdivided according to gender or residence area The prevalence of obesity increased significantly from 7.9% in 1999–2001 to 13.7% in 2009–2010 (p<0.0001), whereas thinness decreased significantly from 10.1% to 2.3% (p<0.0001) in the same periods. The increase of trends in the prevalence of obesity was significantly higher in males (9.7% vs. 17.6%, p = 0.0006) than in females (6.3% vs. 9.8%, p = 0.04) and was slightly higher in urban areas (8.8% vs. 14.3%, p = 0.002) than in rural areas (7.8% vs. 13.0%, p = 0.012). The male gender was associated with a higher risk of being overweight or obese (odds ratio: 1.63; 95% confidence intervals: 1.24–2.15; p = 0.0005) in 2009–2010 than in 1999–2001, after adjusting for age and the residence area. In conclusion, this study showed an increasing trend in the prevalence of overweight and obesity in Sicilian schoolchildren during the last decade and that this trend was related to gender, age and the area of residence. More specifically, our data indicated that the prevalence of obesity increased by 5.8%, the prevalence of thinness decreased by 7.8% and the prevalence of normal-weight children did not change over the course of a decade. These results suggest a shift in the body weights of Sicilian children toward the upper percentiles