Genetic and Chemical Modifiers of a CUG Toxicity Model in Drosophila

Non-coding CUG repeat expansions interfere with the activity of human Muscleblind-like (MBNL) proteins contributing to myotonic dystrophy 1 (DM1). To understand this toxic RNA gain-of-function mechanism we developed a Drosophila model expressing 60 pure and 480 interrupted CUG repeats in the context of a non-translatable RNA. These flies reproduced aspects of the DM1 pathology, most notably nuclear accumulation of CUG transcripts, muscle degeneration, splicing misregulation, and diminished Muscleblind function in vivo. Reduced Muscleblind activity was evident from the sensitivity of CUG-induced phenotypes to a decrease in muscleblind genetic dosage and rescue by MBNL1 expression, and further supported by the co-localization of Muscleblind and CUG repeat RNA in ribonuclear foci. Targeted expression of CUG repeats to the developing eye and brain mushroom bodies was toxic leading to rough eyes and semilethality, respectively. These phenotypes were utilized to identify genetic and chemical modifiers of the CUG-induced toxicity. 15 genetic modifiers of the rough eye phenotype were isolated. These genes identify putative cellular processes unknown to be altered by CUG repeat RNA, and they include mRNA export factor Aly, apoptosis inhibitor Thread, chromatin remodelling factor Nurf-38, and extracellular matrix structural component Viking. Ten chemical compounds suppressed the semilethal phenotype. These compounds significantly improved viability of CUG expressing flies and included non-steroidal anti-inflammatory agents (ketoprofen), muscarinic, cholinergic and histamine receptor inhibitors (orphenadrine), and drugs that can affect sodium and calcium metabolism such as clenbuterol and spironolactone. These findings provide new insights into the DM1 phenotype, and suggest novel candidates for DM1 treatments

Fostering English-taught higher education programs in a Spanish university: the "TechEnglish" innovative project

In recent years, coinciding with adjustments to the Bologna process, many European universities have attempted to improve their international profile by increasing course offerings in English. According to the Institute of International Education (IIE), Spain has notably increased its English-taught higher education programs, ranking fifth in the list of European countries by number of English-taught Master's programs in 2013. This article presents the goals and preliminary results of an on-going innovative education project (TechEnglish) that aims to promote course offerings in English at the Technical University of Madrid (Universidad Politécnica de Madrid, UPM). The UPM is the oldest and largest of all Technical Universities in Spain. It offers graduate and postgraduate programs that cover all the engineering disciplines as well as architecture. Currently, the UPM has no specific bilingual/multilingual program to promote teaching in English, although there is an Educational Model Whitepaper (with a focus on undergraduate degrees) that promotes the development of activities like an International Semester or a unique shared curriculum. The TechEnglish project is an attempt to foster courses taught in English at 7 UPM Technical Schools, including students and 80 faculty members. Four tasks were identified: (1) to design a university wide framework to increase course offerings, (2) to identify administrative difficulties, (3) to increase visibility of courses offered, and (4) to disseminate the results of the project. First, to design a program we analyzed existing programs at other Spanish universities, and other projects and efforts already under way at the UPM. A total of 13 plans were analyzed and classified according to their relation with students (learning), professors (teaching), administration, course offerings, other actors/institutions within the university (e.g., language departments), funds and projects, dissemination activities, mobility plans and quality control. Second, to begin to identify administrative and organizational difficulties in the implementation of teaching in English, we first estimated the current and potential course offerings at the undergraduate level at the UPM using a survey (student, teacher and administrative demand, level of English and willingness to work in English). Third, to make the course offerings more attractive for both Spanish and international students we examined the way the most prestigious universities in Spain and in Europe try to improve the visibility of their academic offerings in English. Finally, to disseminate the results of the project we created a web page and a workspace on the Moodle education platform and prepared conferences and workshops within the UPM. Preliminary results show that increasing course offerings in English is an important step to promote the internationalization of the University. The main difficulties identified at the UPM were related to how to acknowledge/certify the departments, teachers or students involved in English courses, how students should register for the courses, how departments should split and schedule the courses (Spanish and English), and the lack of qualified personnel. A concerted effort could be made to increase the visibility of English-taught programs offered on-line

Six Serum miRNAs Fail to Validate as Myotonic Dystrophy Type 1 Biomarkers

<div><p>Myotonic dystrophy type 1 (DM1) is an autosomal dominant genetic disease caused by expansion of a CTG microsatellite in the 3’ untranslated region of the <i>DMPK</i> gene. Despite characteristic muscular, cardiac, and neuropsychological symptoms, CTG trinucleotide repeats are unstable both in the somatic and germinal lines, making the age of onset, clinical presentation, and disease severity very variable. A molecular biomarker to stratify patients and to follow disease progression is, thus, an unmet medical need. Looking for a novel biomarker, and given that specific miRNAs have been found to be misregulated in DM1 heart and muscle tissues, we profiled the expression of 175 known serum miRNAs in DM1 samples. The differences detected between patients and controls were less than 2.6 fold for all of them and a selection of six candidate miRNAs, <i>miR-103</i>, <i>miR-107</i>, <i>miR-21</i>, <i>miR-29a</i>, <i>miR-30c</i>, and <i>miR-652</i> all failed to show consistent differences in serum expression in subsequent validation experiments.</p></div

Information about the samples used in qPCR.

<p>Information about the samples used in qPCR.</p

Information about the samples used in the miRNA profiling.

<p>Information about the samples used in the miRNA profiling.</p

Validation by q-PCR did not reveal differences in miRNA expression levels between controls and myotonic dystrophy type 1 patients.

<p>(A, B) Graphical representation of the results generated by two algorithms, geNorm and NormFinder, to identify the optimal normalisation miRNA from among all of the candidates (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0150501#pone.0150501.s005" target="_blank">S4 Table</a>). (C) Analysis of the relative expression levels of <i>miR-103</i>, <i>miR-107</i>, <i>miR-21</i>, <i>miR-29a</i>, <i>miR-30c</i>, and <i>miR-652</i> by quantitative PCR on the serum samples of nine DM1 patients and nine healthy individuals. All data were normalised to <i>miR-15</i> expression levels but no significant differences were observed between either group. Graph bars represent average fold-changes of miRNA expression on a logarithmic scale, calculated using the 2^-∆∆Ct method, as well as their confidence intervals. Graph bars represent average fold changes of miRNA expression, calculated using the 2^-∆∆Ct method, along with their standard error.</p

Profiling of miRNA expression levels in myotonic dystrophy type 1 patients and controls.

<p>(A) Heat map graphical representation and clustering analysis of miRNA expression from 9 DM1 patients (P01-P10, excluding P03) and 9 healthy controls (C11-C20 excluding C18). Blue and yellow indicate statistically significant down- and upregulated miRNAs compared to controls, respectively (t-test α  =  0.05). Data is presented as a dendrogram, with the closest branches of the tree showing samples with less dissimilar expression patterns. (B) Statistical analysis of the miRNA profiling carried out with the G*Power tool. These miRNAs have the highest fold-change and Power∼1 statistics in the sample pool. (C) Graphical representation of the expression levels of the miRNAs selected via G*Power analysis. Only <i>miR-21</i> showed a statistically-significant difference when Bonferroni correction was applied. Graph bars represent average fold changes and their standard errors. <i>P</i> > 0.05.</p

The ratio of <i>miR-130a</i> and <i>miR-21</i> failed as a myotonic dystrophy type 1 biomarker.

<p>(A) The ratio of <i>miR-130a</i> and <i>miR-21</i> according to expression levels obtained from the profiling performed with serum samples from nine DM1 patients and nine healthy controls. (B) The same ratio was calculated after measuring <i>miR-130a</i> and <i>miR-21</i> expression levels by quantitative PCR on serum samples from 21 DM1 and 17 control individuals. No statistically-significant differences were observed. Graph bars represent the average ∆Cts (<i>miR-130a</i>--<i>miR-2</i>1) and their standard errors.</p

Quorum sensing network in clinical strains of A. baumannii : AidA is a new quorum quenching enzyme

Acinetobacter baumannii is an important pathogen that causes nosocomial infections generally associated with high mortality and morbidity in Intensive Care Units (ICUs). Currently, little is known about the Quorum Sensing (QS)/Quorum Quenching (QQ) systems of this pathogen. We analyzed these mechanisms in seven clinical isolates of A. baumannii. Microarray analysis of one of these clinical isolates, Ab1 (A. baumannii ST-2-clon-2010), previously cultured in the presence of 3-oxo-C12-HSL (a QS signalling molecule) revealed a putative QQ enzyme (α/β hydrolase gene, AidA). This QQ enzyme was present in all nonmotile clinical isolates (67% of which were isolated from the respiratory tract) cultured in nutrient depleted LB medium. Interestingly, this gene was not located in the genome of the only motile clinical strain growing in this medium (A. baumannii strain Ab421-GEIH-2010 [Ab7], isolated from a blood sample). The AidA protein expressed in E. coli showed QQ activity. Finally, we observed downregulation of the AidA protein (QQ system attenuation) in the presence of HO (ROS stress). In conclusion, most of the A. baumannii clinical strains were not surface motile (84%) and were of respiratory origin (67%). Only the pilT gene was involved in surface motility and related to the QS system. Finally, a new QQ enzyme (α/β hydrolase gene, AidA protein) was detected in these strains