Mechanistic basis of an epistatic interaction reducing age at onset in hereditary spastic paraplegia

Many genetic neurological disorders exhibit variable expression within affected families, often exemplified by variations in disease age at onset. Epistatic effects (i.e. effects of modifier genes on the disease gene) may underlie this variation, but the mechanistic basis for such epistatic interactions is rarely understood. Here we report a novel epistatic interaction between SPAST and the contiguous gene DPY30, which modifies age at onset in hereditary spastic paraplegia, a genetic axonopathy. We found that patients with hereditary spastic paraplegia caused by genomic deletions of SPAST that extended into DPY30 had a significantly younger age at onset. We show that, like spastin, the protein encoded by SPAST, the DPY30 protein controls endosomal tubule fission, traffic of mannose 6-phosphate receptors from endosomes to the Golgi, and lysosomal ultrastructural morphology. We propose that additive effects on this pathway explain the reduced age at onset of hereditary spastic paraplegia in patients who are haploinsufficient for both genes.This work was supported by grants to E.R.; Project Grant from United States Spastic Paraplegia Foundation, UK Medical Research Council Project Grant [MR/M00046X/1], Project grant from NIHR Biomedical Research Centre at Addenbrooke’s Hospital, Wellcome Trust Senior Research Fellowship in Clinical Science [082381], Project Grant from Tom Wahlig Stiftung (project 33). J.E. and P.M. are supported by a Wellcome Trust Principal Research Fellowship Grant to Margaret S. Robinson [086598]. T.M.N. was supported by an MRC PhD studentship [G0800117]. B.W. is supported by the Tom Wahlig Advanced Fellowship, the German Federal Ministry of Education and Research (BMBF, 01GQ113), the Bavarian Ministry of Education and Culture, Sciences and Arts in the framework of the Bavarian Molecular Biosystems Research Network and ForIPS, and the Interdisciplinary Centre for Clinical Research (IZKF, University Hospital of Erlangen, N3 and F3). T.R. was supported by research grant DFG GRK2162/1 of the Deutsche Forschungsgemeinschaft. The study was also supported by the European Union within the 7th European Community Framework Program for Research and Technological Development through funding for the NEUROMICS network (F5-2012-305121 to L.S. and A.D.), the E-Rare Network NEUROLIPID (01GM1408B to R.S. and ANR-13-RARE-0003-02 to G.S.), and a Marie Curie International Outgoing Fellowship (grant PIOF-GA-2012-326681 to R.S. and L.S.). This work was further supported by the US National Institutes of Health (NIH) (grant 5R01NS072248 to R.S.), the German HSP-Selbsthilfegruppe e.V. (grant to R.S. and L.S.), and grants to C.B.: Project Grant from Tom Wahlig Stiftung (project 20), grant from the Stiftung für Pathobiochemie und Molekulare Diagnostik. CIMR is supported by a Wellcome Trust Strategic Award [100140] and Equipment Grant [093026]

Does inequality erode generalized trust? Evidence from Romanian youths

Generalized trust is a critical component of liberal democratic citizenship. We evaluate the extent to which exposure to socioeconomic inequality erodes trust among Romanian youths. Using national survey data of Romanian eighth-grade and high school students, we evaluate this effect as a product of socioeconomic diversity within the classroom, controlling for the social status of the students as well as socioeconomic inequality within the community where the school is located. Our analysis shows that generalized trust is higher for students in higher grades. However, despite this maturing effect, students exposed to greater levels of socioeconomic diversity have significantly lower levels of trust. The effect is particularly acute for students in the ninth grade. This finding holds when controlling for socioeconomic diversity and polarization in the community. The result reinforces the idea that generalized trust develops early in one’s life and is quite stable, although a major life transformation, such as entering high school, may alter trust depending on the social context

Cross-sectional study of availability and pharmaceutical quality of antibiotics requested with or without prescription (Over The Counter) in Surabaya, Indonesia

Contains fulltext : 87881.pdf (publisher's version ) (Open Access

Neoplastic transformation of breast epithelial cells by genotoxic stress

<p>Abstract</p> <p>Background</p> <p>Exposure to genotoxic stresses such as radiation and tobacco smoke can cause increased cancer incidence rate as reflected in an in depth meta-analysis of data for women and breast cancer incidence. Published reports have indicated that exposures to low dose radiation and tobacco smoke are factors that contribute to the development of breast cancer. However, there is a scarcity of information on the combinatorial effects of low dose radiation and tobacco smoke on formation and progression of breast cancer. The combination of these two genotoxic insults can induce significant damage to the genetic material of the cells resulting in neoplastic transformation.</p> <p>Methods</p> <p>To study the effects of low dose ionizing radiation and tobacco smoke on breast cells, MCF 10A cells were treated either with radiation (Rad - 0.1 Gray) or cigarette smoke condensate (Csc - 10 microgram/ml of medium) or a combination of Rad + Csc. Following treatments, cells were analyzed for cell cycle distribution patterns and the ability to extrude the Hoechst 33342 dye. In addition, <it>in vitro </it>invasion and migration as well as mammosphere formation assays were performed. Finally, differential gene expression profiles were generated from the individual and combination treatment.</p> <p>Results</p> <p>Exposure of MCF 10A cells to the combination of radiation plus cigarette smoke condensate generated a neoplastic phenotype. The transformed phenotype promoted increased mammosphere numbers, altered cell cycle phases with a doubling of the population in S phase, and increased invasion and motility. Also, exclusion of Hoechst 33342 dye, a surrogate marker for increased ABC transporters, was observed, which indicates a possible increase in drug resistance. In addition, changes in gene expression include the up regulation of genes encoding proteins involved in metabolic pathways and inflammation.</p> <p>Conclusions</p> <p>The results indicate that when normal breast cells are exposed to low dose radiation in combination with cigarette smoke condensate a phenotype is generated that exhibits traits indicative of neoplastic transformation. More importantly, this is the first study to provide a new insight into a possible etiology for breast cancer formation in individuals exposed to low dose radiation and tobacco smoke.</p

Coordinate up-regulation of TMEM97 and cholesterol biosynthesis genes in normal ovarian surface epithelial cells treated with progesterone: implications for pathogenesis of ovarian cancer

<p>Abstract</p> <p>Background</p> <p>Ovarian cancer (OvCa) most often derives from ovarian surface epithelial (OSE) cells. Several lines of evidence strongly suggest that increased exposure to progesterone (P4) protects women against developing OvCa. However, the underlying mechanisms of this protection are incompletely understood.</p> <p>Methods</p> <p>To determine downstream gene targets of P4, we established short term <it>in vitro </it>cultures of non-neoplastic OSE cells from six subjects, exposed the cells to P4 (10^-6M) for five days and performed transcriptional profiling with oligonucleotide microarrays containing over 22,000 transcripts.</p> <p>Results</p> <p>We identified concordant but modest gene expression changes in cholesterol/lipid homeostasis genes in three of six samples (responders), whereas the other three samples (non-responders) showed no expressional response to P4. The most up-regulated gene was <it>TMEM97 </it>which encodes a transmembrane protein of unknown function (MAC30). Analyses of outlier transcripts, whose expression levels changed most significantly upon P4 exposure, uncovered coordinate up-regulation of 14 cholesterol biosynthesis enzymes, insulin-induced gene 1, low density lipoprotein receptor, <it>ABCG1</it>, endothelial lipase, stearoyl- CoA and fatty acid desaturases, long-chain fatty-acyl elongase, and down-regulation of steroidogenic acute regulatory protein and <it>ABCC6</it>. Highly correlated tissue-specific expression patterns of <it>TMEM97 </it>and the cholesterol biosynthesis genes were confirmed by analysis of the GNF Atlas 2 universal gene expression database. Real-time quantitative RT-PCR analyses revealed 2.4-fold suppression of the <it>TMEM97 </it>gene expression in short-term cultures of OvCa relative to the normal OSE cells.</p> <p>Conclusion</p> <p>These findings suggest that a co-regulated transcript network of cholesterol/lipid homeostasis genes and <it>TMEM97 </it>are downstream targets of P4 in normal OSE cells and that <it>TMEM97 </it>plays a role in cholesterol and lipid metabolism. The P4-induced alterations in cholesterol and lipid metabolism in OSE cells might play a role in conferring protection against OvCa.</p

Does the perception of fairness and standard of care in the health system depend on the field of study? Results of an empirical analysis

Background: The main challenge in the context of health care reforms and priority setting is the establishment and/or maintenance of fairness and standard of care. For the political process and interdisciplinary discussion, the subjective perception of the health care system might even be as important as potential objective criteria. Of special interest are the perceptions of academic disciplines, whose representatives act as decision makers in the health care sector. The aim of this study is to explore and compare the subjective perception of fairness and standard of care in the German health care system among students of medicine, law, economics, philosophy, and religion. Methods: Between October 2011 and January 2012, we asked freshmen and advanced students of the fields mentioned above to participate in a paper and pencil survey. Prior to this, we formulated hypotheses. The data were analysed by micro econometric regression techniques. Results: Data from 1,088 students were included in the study. Medical students, freshmen, and advanced students perceive the standard of care significantly as being better than non-medical students. Differences in the perception of fairness are not significant between the freshmen of the academic disciplines; however, they increase with the number of study terms. Besides the field of study, further variables such as gender and health status have a significant impact on perceptions. Conclusions: Our results show that there are differences in the perception of fairness and standard of care between academic disciplines, which might influence the interdisciplinary discussion on health care reforms and priority setting.Leibniz University Hannover/Wege in die Forschung I

Methods to Study Centrosomes and Cilia in Drosophila

The deposited item is a book chapter and is part of the series " Methods in Molecular Biology book series ([MIMB, volume 1454]) published by the publisher Humana Press.The deposited book chapter is a pre-print version and hasn't been submitted to peer reviewing.There is no public supplementary material available for this publication.This publication hasn't any creative commons license associated.Centrioles and cilia are highly conserved eukaryotic organelles. Drosophila melanogaster is a powerful genetic and cell biology model organism, extensively used to discover underlying mechanisms of centrosome and cilia biogenesis and function. Defects in centrosomes and cilia reduce fertility and affect different sensory functions, such as proprioception, olfaction, and hearing. The fly possesses a large diversity of ciliary structures and assembly modes, such as motile, immotile, and intraflagellar transport (IFT)-independent or IFT-dependent assembly. Moreover, all the diverse ciliated cells harbor centrioles at the base of the cilia, called basal bodies, making the fly an attractive model to better understand the biology of this organelle. This chapter describes protocols to visualize centrosomes and cilia by fluorescence and electron microscopy.Fundação Portuguesa para a Ciência e Tecnologia grants: (SFRH/BPD/87479/2012, SFRH/BD/52176/2013); EMBO installation grant; ERC starting grant.info:eu-repo/semantics/publishedVersio

Membrane Docking Geometry of GRP1 PH Domain Bound to a Target Lipid Bilayer: An EPR Site-Directed Spin-Labeling and Relaxation Study

The second messenger lipid PIP3 (phosphatidylinositol-3,4,5-trisphosphate) is generated by the lipid kinase PI3K (phosphoinositide-3-kinase) in the inner leaflet of the plasma membrane, where it regulates a broad array of cell processes by recruiting multiple signaling proteins containing PIP3-specific pleckstrin homology (PH) domains to the membrane surface. Despite the broad importance of PIP3-specific PH domains, the membrane docking geometry of a PH domain bound to its target PIP3 lipid on a bilayer surface has not yet been experimentally determined. The present study employs EPR site-directed spin labeling and relaxation methods to elucidate the membrane docking geometry of GRP1 PH domain bound to bilayer-embedded PIP3. The model target bilayer contains the neutral background lipid PC and both essential targeting lipids: (i) PIP3 target lipid that provides specificity and affinity, and (ii) PS facilitator lipid that enhances the PIP3 on-rate via an electrostatic search mechanism. The EPR approach measures membrane depth parameters for 18 function-retaining spin labels coupled to the PH domain, and for calibration spin labels coupled to phospholipids. The resulting depth parameters, together with the known high resolution structure of the co-complex between GRP1 PH domain and the PIP3 headgroup, provide sufficient constraints to define an optimized, self-consistent membrane docking geometry. In this optimized geometry the PH domain engulfs the PIP3 headgroup with minimal bilayer penetration, yielding the shallowest membrane position yet described for a lipid binding domain. This binding interaction displaces the PIP3 headgroup from its lowest energy position and orientation in the bilayer, but the headgroup remains within its energetically accessible depth and angular ranges. Finally, the optimized docking geometry explains previous biophysical findings including mutations observed to disrupt membrane binding, and the rapid lateral diffusion observed for PIP3-bound GRP1 PH domain on supported lipid bilayers

Imaginal Discs – A New Source of Chromosomes for Genome Mapping of the Yellow Fever Mosquito Aedes aegypti

Dengue fever is an emerging health threat to as much as half of the human population around the world. No vaccines or drug treatments are currently available. Thus, disease prevention is largely based on efforts to control its major mosquito vector Ae. aegypti. Novel vector control strategies, such as population replacement with pathogen-incompetent transgenic mosquitoes, rely on detailed knowledge of the genome organization for the mosquito. However, the current genome assembly of Ae. aegypti is highly fragmented and requires additional physical mapping onto chromosomes. The absence of readable polytene chromosomes makes genome mapping for this mosquito extremely challenging. In this study, we discovered and investigated a new source of chromosomes useful for the cytogenetic analysis in Ae. aegypti – mitotic chromosomes from imaginal discs of 4th instar larvae. Using natural banding patterns of these chromosomes, we developed a new band-based approach for physical mapping of DNA probes to the precise chromosomal positions. Further application of this approach for genome mapping will greatly enhance the utility of the existing draft genome sequence assembly for Ae. aegypti and thereby facilitate application of advanced genome technologies for investigating and developing novel genetic control strategies for dengue transmission

Model-Based Deconvolution of Cell Cycle Time-Series Data Reveals Gene Expression Details at High Resolution

In both prokaryotic and eukaryotic cells, gene expression is regulated across the cell cycle to ensure “just-in-time” assembly of select cellular structures and molecular machines. However, present in all time-series gene expression measurements is variability that arises from both systematic error in the cell synchrony process and variance in the timing of cell division at the level of the single cell. Thus, gene or protein expression data collected from a population of synchronized cells is an inaccurate measure of what occurs in the average single-cell across a cell cycle. Here, we present a general computational method to extract “single-cell”-like information from population-level time-series expression data. This method removes the effects of 1) variance in growth rate and 2) variance in the physiological and developmental state of the cell. Moreover, this method represents an advance in the deconvolution of molecular expression data in its flexibility, minimal assumptions, and the use of a cross-validation analysis to determine the appropriate level of regularization. Applying our deconvolution algorithm to cell cycle gene expression data from the dimorphic bacterium Caulobacter crescentus, we recovered critical features of cell cycle regulation in essential genes, including ctrA and ftsZ, that were obscured in population-based measurements. In doing so, we highlight the problem with using population data alone to decipher cellular regulatory mechanisms and demonstrate how our deconvolution algorithm can be applied to produce a more realistic picture of temporal regulation in a cell