New stably transfected bioluminescent cells expressing FLAG epitope-tagged estrogen receptors to study their chromatin recruitment

International audienceBACKGROUND: Biological actions of estrogens are mediated by the two specific estrogen receptors ERalpha and ERbeta. However, due to the absence of adequate cellular models, their respective transcriptional activities are still poorly understood. For instance, the evaluation of such differing properties on the transcription of responsive genes using ChIP experiments was hindered by the deficiency of cells exhibiting the same genotypic background and properties but expressing only one of the ERs. We describe here the generation of such cells, using an estrogen receptor negative HELN cell line that was derived from HeLa cells stably transfected with an ERE-driven luciferase plasmid. These HELN-Falpha and HELN-Fbeta cell lines stably express either the alpha or beta (full length) estrogen receptor tagged with the FLAG epitope. The use of antibodies directed against the FLAG epitope allowed a direct comparative evaluation of the respective actions of both ERs using ChIP. RESULTS: HELN-Falpha and HELN-Fbeta cell lines were found to express comparable levels of their corresponding tagged receptors with a Kd for estradiol binding of 0.03 and 0.27 nM respectively. The presence of a stably transfected ERE-driven luciferase plasmid in these cells allowed the direct evaluation of the transcriptional activity of both tagged receptors, using natural or synthetic estrogens. FLAG-ERalpha and FLAG-ERbeta were found to exhibit similar transcriptional activity, as indicated by a kinetic evaluation of the transcriptional activation of the luciferase gene during 10 hrs of treatment with estradiol. The validity of these model cells was further confirmed by the predictable transcriptional regulations measured upon treatments with ERalpha or ERbeta specific ligands. The similar immunoprecipitation efficiency of both tagged receptors by an anti-FLAG antibody allowed the assessment of their kinetic recruitment on the synthetic luciferase promoter (containing an estrogen response element) by ChIP assays during 8 hours. A biphasic curve was obtained for both FLAG-ERalpha and FLAG-ERbeta, with a peak occurring either at 2 hr or at 1 hr, respectively, and a second one following 4 hr of E2 stimulation in both cases. In MCF-7 cells, the recruitment of ERalpha also exhibited a biphasic behaviour; with the second peak however not so important than in the HeLa cell lines. CONCLUSION: In HELN derived cell lines, no fundamental differences between kinetics were observed during 8 hours for FLAG-ERalpha and FLAG-ERbeta, as well as for polymerase II recruitment. However, the relative importance of recruitment between 1 hr and 4 hr was found to be different in HeLa cell line expressing exogenous tagged ERalpha and in MCF-7 cell line expressing endogenous ER

A generic finite element framework on parallel tree-based adaptive meshes

We present highly scalable parallel distributed-memory algorithms and associated data structures for a generic finite element framework that supports h-adaptivity on computational domains represented as multiple connected adaptive trees—forest-of-trees—, thus providing multi-scale resolution on problems governed by partial differential equations.The framework is grounded on a rich representation of the adaptive mesh suitable for generic finite elements that is built on top of a low-level, light-weight forest-oftrees data structure handled by a specialized, highly parallel adaptive meshing engine. Along the way, we have identified the requirements that the forest-of-trees layer must fulfill to be coupled into our framework. Essentially, it must be able to describe neighboring relationships between cells in the adapted mesh (apart from hierarchical relationships) across the lower-dimensional objects at the boundary of the cells. Atop this two-layered mesh representation, we build the rest of data structures required for the numerical integration and assembly of the discrete system of linear equations.We consider algorithms that are suitable for both subassembled and fully-assembled distributed data layouts of linear system matrices. The proposed framework has been implemented within the FEMPAR scientific software library, using p4est as a practical forest-of-octrees demonstrator. A comprehensive strong scaling study of this implementation when applied to Poisson and Maxwell problems reveals remarkable scalability up to 32.2K CPU cores and 482.2M degrees of freedom. Besides, the implementation in FEMPAR of the proposed approach is up to 2.6 and 3.4 times faster than the state-of-the-art deal.II finite element software in the h-adaptive approximation of a Poisson problem with firstand second-order Lagrangian finite elements, respectively (excluding the linear solver step from the comparison)

Numerical modelling of heat transfer and experimental validation in Powder-Bed Fusion with the Virtual Domain Approximation

Among metal additive manufacturing technologies, powder-bed fusion features very thin layers and rapid solidification rates, leading to long build jobs and a highly localized process. Many efforts are being devoted to accelerate simulation times for practical industrial applications. The new approach suggested here, the virtual domain approximation, is a physics-based rationale for spatial reduction of the domain in the thermal finite-element analysis at the part scale. Computational experiments address, among others, validation against a large physical experiment of 17.5 $\mathrm{[cm^3]}$ of deposited volume in 647 layers. For fast and automatic parameter estimation at such level of complexity, a high-performance computing framework is employed. It couples FEMPAR-AM, a specialized parallel finite-element software, with Dakota, for the parametric exploration. Compared to previous state-of-the-art, this formulation provides higher accuracy at the same computational cost. This sets the path to a fully virtualized model, considering an upwards-moving domain covering the last printed layers

The nuclear receptor transcriptional coregulator RIP140

The nuclear receptor superfamily comprises ligand-regulated transcription factors that control various developmental and physiological pathways. These receptors share a common modular structure and regulate gene expression through the recruitment of a large set of coregulatory proteins. These transcription cofactors regulate, either positively or negatively, chromatin structure and transcription initiation. One of the first proteins to be identified as a hormone-recruited cofactor was RIP140. Despite its recruitment by agonist-liganded receptors, RIP140 exhibits a strong transcriptional repressive activity which involves several inhibitory domains and different effectors. Interestingly, the RIP140 gene, located on chromosome 21 in humans, is finely regulated at the transcriptional level by various nuclear receptors. In addition, the protein undergoes several post-translational modifications which control its repressive activity. Finally, experiments performed in mice devoid of the RIP140 gene indicate that this transcriptional cofactor is essential for female fertility and energy homeostasis. RIP140 therefore appears to be an important modulator of nuclear receptor activity which could play major roles in physiological processes and hormone-dependent diseases

An artificial moth: Chemical source localization using a robot based neuronal model of moth optomotor anemotactic search

Robots have been used to model nature, while nature in turn can contribute to the real-world artifacts we construct. One particular domain of interest is chemical search where a number of efforts are underway to construct mobile chemical search and localization systems. We report on a project that aims at constructing such a system based on our understanding of the pheromone communication system of the moth. Based on an overview of the peripheral processing of chemical cues by the moth and its role in the organization of behavior we emphasize the multimodal aspects of chemical search, i.e. optomotor anemotactic chemical search. We present a model of this behavior that we test in combination with a novel thin metal oxide sensor and custom build mobile robots. We show that the sensor is able to detect the odor cue, ethanol, under varying flow conditions. Subsequently we show that the standard model of insect chemical search, consisting of a surge and cast phases, provides for robust search and localization performance. The same holds when it is augmented with an optomotor collision avoidance model based on the Lobula Giant Movement Detector (LGMD) neuron of the locust. We compare our results to others who have used the moth as inspiration for the construction of odor robot

The Aging Males' Symptoms (AMS) scale: review of its methodological characteristics

BACKGROUND: The current paper reviews data from different sources to get a closer impression on the psychometric and other methodological characteristics of the Aging Males' Symptoms (AMS) scale gathered recently. The scale was designed and standardized as self-administered scale to (a) to assess symptoms of aging (independent from those which are disease-related) between groups of males under different conditions, (b) to evaluate the severity of symptoms over time, and (c) to measure changes pre- and post androgen replacement therapy. The scale is in widespread use (14 languages). METHOD: Original data from different studies in many countries were centrally analysed to evaluate reliability and validity of the AMS. RESULTS: Reliability measures (consistency and test-retest stability) were found to be good across countries, although the sample size was sometimes small. Validity: The internal structure of the AMS in healthy and androgen deficient males, and across countries was sufficiently similar to conclude that the scale really measures the same phenomenon. The sub-scores and total score correlations were high (0.8–0.9) but lower among the sub-scales (0.5–0.7). This however suggests that the subscales are not fully independent. The comparison with other scales for aging males or screening instruments for androgen deficiency showed sufficiently good correlations, illustrating a good criterion-oriented validity. The same is true for the comparison with the generic quality-of-life scale SF36 where also high correlation coefficients have been shown. Methodological analyses of a treatment study of symptomatic males with testosterone demonstrated the ability of the AMS scale to measure treatment effect, irrespective of the severity of complaints before therapy. It was also shown that the AMS result can predict the independently generated (physician's) opinion about the individual treatment effect. CONCLUSION: The currently available methodological evidence points towards a high quality of the AMS scale to measure and to compare HRQoL of aging males over time or before/after treatment, it suggests a high reliability and high validity as far as the process of construct validation could be pressed ahead yet. But certainly more data will become available, particularly from ongoing clinical studies

The Aging Males' Symptoms (AMS) scale: Update and compilation of international versions

BACKGROUND: The interest of clinical research in aging males increased in recent years and thereby the interest to measure health-related quality of life (HRQoL) and symptoms of aging men. The Aging Males' Symptoms scale (AMS) became the most commonly used scale to measure HRQoL and symptoms in aging males in many countries worldwide. The aim of this paper is to review the current state of the instrument particularly concerning versions of the scale in different languages in the light of the quality of the translation process. AMS VERSIONS AVAILABLE: Most of the translations were performed following international methodological recommendations for linguistic & cultural adaptation of HRQoL instruments. Mainly the English version was used as source language for the translation into Dutch, Spanish, Portuguese, Italian, Swedish, and Japanese (attached as additional PDF-files). Preliminary versions that were derived only from forward translations are of secondary quality and available in Finnish, Flemish, and Russian. It is recommended to complete the translation process for the latter languages before using them in international studies. TRANSLATIONS IN PROCESS: The AMS scale is in the process of consensus finding of two existing French versions, and the versions in the Korean, Thai, and Indonesian languages have not yet been completed in the translation process. CONCLUSION: The AMS scale is obviously a valuable tool for assessing health related quality of life in aging men, because it is used worldwide. It is a standardized scale according to psychometric norms. Most of the currently available language versions were translated following international standards for linguistic and cultural translation of quality of life scales. Assistance is offered to help interested parties in the translation process