Genetic Rescue of Fragile X Syndrome Links FMRP Deficiency to Codon Optimality-Dependent RNA Destabilization [preprint]

Fragile X syndrome (FXS) is caused by inactivation of FMR1 gene and loss of its encoded product the RNA binding protein FMRP, which generally represses translation of its target transcripts in the brain. In mouse models of FXS (i.e., Fmr1 knockout animals; Fmr1 KO), deletion of Cpeb1, which encodes a translational activator, mitigates nearly all pathophysiologies associated with the disorder. Here we reveal unexpected wide-spread dys-regulation of RNA abundance in Fmr1 KO brain cortex and its rescue to normal levels in Fmr1/Cpeb1 double KO mice. Alteration and restoration of RNA levels are the dominant molecular events that drive the observed dys-regulation and rescue of translation as measured by whole transcriptome ribosome occupany in the brain. The RNAs down-regulated and rescued in these animal models are highly enriched for FMRP binding targets and have an optimal codon bias that would predict their stability in wild type and possible instability in FMRP knock-out brain. Indeed, whole transcriptome analysis of RNA metabolic rates demonstrates a codon optimality-dependent elevation of RNA destruction in FMRP knock-out cortical neurons. This elevated RNA destruction leads to a massive reshuffling of the identities of stabilizing versus destabilizing codons in neurons upon loss of FMRP. Our results show a widespread RNA instability in FXS, which results from the uncoupling of codon optimality, ribosome occupancy, and RNA degradation mechanisms. Re-establishment of the linkage among these events is likely required by the genetic rescue of the disorder

FMRP Links Optimal Codons to mRNA stability in Neurons [preprint]

Fragile X syndrome (FXS) is caused by inactivation of the FMR1 gene and loss of encoded FMRP, an RNA binding protein that represses translation of some of its target transcripts. Here we use ribosome profiling and RNA-seq to investigate the dysregulation of translation in the mouse brain cortex. We find that most changes in ribosome occupancy on hundreds of mRNAs are largely driven by dysregulation in transcript abundance. Many downregulated mRNAs, which are mostly responsible for neuronal and synaptic functions, are highly enriched for FMRP binding targets. RNA metabolic labeling demonstrates that in FMRP-deficient cortical neurons, mRNA downregulation is caused by elevated degradation, and is correlated with codon optimality. Moreover, FMRP preferentially binds mRNAs with optimal codons, suggesting that it stabilizes such transcripts through direct interactions via the translational machinery. Finally, we show that the paradigm of genetic rescue of FXS-like phenotypes in FMRP-deficient mice by deletion of the Cpeb1 gene is mediated by restoration of steady state RNA levels and consequent rebalancing of translational homeostasis. Our data establish an essential role of FMRP in codon optimality-dependent mRNA stability as an important factor in FXS

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Qualität von Lehrkräftefortbildungen einschätzen. Ein Arbeitsbuch aus dem Projekt IMPRESS

Die Erhaltung und Weiterentwicklung der eigenen Kompetenzen ist neben dem Unterrichten ein wichtiger Bestandteil der beruflichen Tätigkeiten von Lehrpersonen. Für die Nutzung von formalen Fortbildungen können positive Wirkungen auf die professionellen Kompetenzen von Lehrpersonen, die Qualität des Unterrichts und das Lernen von Schüler:innen nachgewiesen werden. Die Forschung zur Fort- und Weiterbildung von Lehrkräften konzentrierte sich bisher weitgehend darauf, die inhaltlichen und strukturellen Merkmale wirksamer Angebote zu identifizieren und zu erfassen, während die Lehr-Lern-Prozesse der durchgeführten Fortbildungen noch weitgehend unberücksichtigt blieben. Die Universitäten Kassel, Jena und Potsdam haben sich in dem gemeinsamen Projekt „InstruMent zur Erfassung der PRozESSqualität von Lehrkräftefortbildungen“ (IMPRESS) zum Ziel gesetzt, ein Instrument zu entwickeln und bereit zu stellen, mit dem die Qualität von Lehr-Lern-Prozessen in Fortbildungen erfasst und daraufhin besser beschrieben werden kann. Das Projekt IMPRESS greift damit ein Desiderat auf und leistet einen wichtigen Beitrag zur Erforschung der Qualität von Lehrkräftefortbildungen. Das vorliegende Arbeitsbuch stellt zunächst Hintergründe zu den Konstrukten der Prozessqualität dar und dokumentiert anschließend die neu entwickelten Erhebungsinstrumente. Es handelt sich um Befragungsinstrumente für die Fortbildungsteilnehmer:innen und die Fortbildner:innen sowie um ein Instrument für die externe Beobachtung. Die Qualität der Lehr-Lern-Prozesse wird in den Instrumenten durch insgesamt zehn Dimensionen konzeptualisiert und operationalisiert. Darüber hinaus werden in dem Arbeitsbuch zwei weitere Instrumente zur Verfügung gestellt, um sowohl inhaltliche und strukturelle Merkmale einer Fortbildung als auch das Auftreten und Handeln von Fortbildner:innen aus der Perspektive der teilnehmenden Lehrpersonen zu erfassen. (Autor*innen)Maintaining and developing one\u27s own competencies, besides teaching, is an important part of the professional activities of teachers. The use of formal professional development programs has been proven to have positive effects on teachers\u27 professional competencies, the quality of teaching, and student learning. Research on teacher professional development has thus focused mainly on identifying and assessing the content-related and structural characteristics of effective professional development programs, while teaching-learning processes of teacher professional development has hardly been investigated. The Universities of Kassel, Jena and Potsdam collaborated on the project "Survey for the assessment of the quality of teaching-learning-processes in teacher professional development" (IMPRESS), which aims to develop and provide an instrument for assessing the quality of teaching-learning processes in professional development programs. The IMPRESS project thus addresses a desideratum and makes an important contribution to research on teachers\u27 professional development. This workbook first provides a theoretical grounding on the constructs of process quality and then documents the newly developed instruments. These are questionnaires for the participants and the facilitators as well as an instrument for external observation. The quality of the teaching-learning processes is conceptualized and operationalized in the instruments by a total of ten dimensions. In addition, the workbook provides two instruments to measure both the content and structural features of a professional development program as well as the appearance and behavior of the facilitators from the perspective of the participating teachers. (Authors

Local Optical Spectroscopy in Quantum Confined Systems: A Theoretical Description

A theoretical description of local absorption is proposed in order to investigate spectral variations on a length scale comparable with the extension of the relevant quantum states. A general formulation is derived within the density-matrix formalism including Coulomb correlation, and applied to the prototypical case of coupled quantum wires. The results show that excitonic effects may have a crucial impact on the local absorption with implications for the spatial resolution and the interpretation of near-field optical spectra.Comment: To appear in Phys. Rev. Lett. - 11 pages, 3 PostScript figures (1 figure in colors) embedded. Uses RevTex, and psfig style

Geometrically defined environments direct cell division rate and subcellular YAP localization in single mouse embryonic stem cells

Mechanotransduction via yes-associated protein (YAP) is a central mechanism for decision-making in mouse embryonic stem cells (mESCs). Nuclear localization of YAP is tightly connected to pluripotency and increases the cell division rate (CDR). How the geometry of the extracellular environment influences mechanotransduction, thereby YAP localization, and decision-making of single isolated mESCs is largely unknown. To investigate this relation, we produced well-defined 2D and 2.5D microenvironments and monitored CDR and subcellular YAP localization in single mESCs hence excluding cell–cell interactions. By systematically varying size and shape of the 2D and 2.5D substrates we observed that the geometry of the growth environment affects the CDR. Whereas CDR increases with increasing adhesive area in 2D, CDR is highest in small 2.5D micro-wells. Here, mESCs attach to all four walls and exhibit a cross-shaped cell and nuclear morphology. This observation indicates that changes in cell shape are linked to a high CDR. Inhibition of actomyosin activity abrogate these effects. Correspondingly, nuclear YAP localization decreases in inhibitor treated cells, suggesting a relation between cell shape, intracellular forces, and cell division rate. The simplicity of our system guarantees high standardization and reproducibility for monitoring stem cell reactions and allows addressing a variety of fundamental biological questions on a single cell level

Ultrahard carbon film from epitaxial two-layer graphene

Atomically thin graphene exhibits fascinating mechanical properties, although its hardness and transverse stiffness are inferior to those of diamond. To date, there hasn't been any practical demonstration of the transformation of multi-layer graphene into diamond-like ultra-hard structures. Here we show that at room temperature and after nano-indentation, two-layer graphene on SiC(0001) exhibits a transverse stiffness and hardness comparable to diamond, resisting to perforation with a diamond indenter, and showing a reversible drop in electrical conductivity upon indentation. Density functional theory calculations suggest that upon compression, the two-layer graphene film transforms into a diamond-like film, producing both elastic deformations and sp2-to-sp3 chemical changes. Experiments and calculations show that this reversible phase change is not observed for a single buffer layer on SiC or graphene films thicker than 3 to 5 layers. Indeed, calculations show that whereas in two-layer graphene layer-stacking configuration controls the conformation of the diamond-like film, in a multilayer film it hinders the phase transformation.Comment: Published online on Nature Nanotechnology on December 18, 201

HE4 as a serum biomarker for the diagnosis of pelvic masses: a prospective, multicenter study in 965 patients

Background: To evaluate the diagnostic value of adding human epididymis protein 4 (HE4), cancer antigen 125 (CA125) and risk of malignancy algorithm (ROMA) to ultrasound for detecting ovarian cancer in patients with a pelvic mass. Methods: This was a prospective, observational, multicenter study. Patients aged > 18 years who were scheduled to undergo surgery for a suspicious pelvic mass had CA125 and HE4 levels measured prior to surgery, in addition to a routine transvaginal ultrasound scan. The diagnostic performance of CA125, HE4 and ROMA for distinguishing between benign and malignant adnexal masses was assessed using receiver operating characteristic (ROC) analysis and the corresponding area under the curve (AUC). Results: Of 965 evaluable patients, 804 were diagnosed with benign tumors and 161 were diagnosed with ovarian cancer. In late-stage ovarian cancer, CA125, HE4 and ROMA all had an excellent diagnostic performance (AUC > 0.92), whereas in stage I and II, diagnostic performance of all three biomarkers was less adequate (AUC < 0.77). In the differential diagnosis of ovarian cancer and endometriosis, ROMA and HE4 performed better than CA125 with 99 and 98.1% versus 75.0% sensitivity, respectively, at 75.4% specificity. Conclusions: ROMA and HE4 could be valuable biomarkers to help with the diagnosis of ovarian cancer in premenopausal patients in order to differentiate from endometriosis, whereas CA125 may be more adequate for postmenopausal patients

Enhanced Biological Activity of BMP‐2 Bound to Surface‐Grafted Heparan Sulfate

Over the last decade, there has been a growing interest in the development of new materials to improve bone morphogenetic protein‐2 (BMP‐2) delivery for tissue regeneration. This study reports the development and application of model surfaces that present BMP‐2 via heparan sulfate (HS), a ubiquitous component of the extracellular matrix (ECM). On these surfaces, HS is grafted by its reducing end, to mimic the natural arrangement of HS proteoglycans in the ECM. The binding of each component on these biomimetic surfaces is highly controlled, in terms of stoichiometry of molecules and BMP‐2/grafted‐HS affinity, as determined by surface‐sensitive techniques. For comparison, this study also uses surfaces presenting immobilized BMP‐2 alone. Functional validations of the surfaces are performed using a murine myoblast cell line (C2C12) and primary human mesenchymal stromal cells. In both cell types, HS‐bound BMP‐2 and surface‐immobilized BMP‐2 significantly prolong SMAD 1/5 phosphorylation, compared to BMP‐2 added to the culture media. Moreover, HS‐bound BMP‐2 enhances p‐SMAD 1/5 levels in C2C12 cells and reduces noggin antagonistic activity. Thus, grafted HS positively affects BMP‐2 cellular activity. This innovative surface design, which mimics natural interactions of growth factors with ECM components, constitutes a promising candidate for future regenerative medicine applications