Unraveling the regulation of mTORC2 using logical modeling

Background The mammalian target of rapamycin (mTOR) is a regulator of cell proliferation, cell growth and apoptosis working through two distinct complexes: mTORC1 and mTORC2. Although much is known about the activation and inactivation of mTORC1, the processes controlling mTORC2 remain poorly characterized. Experimental and modeling studies have attempted to explain the regulation of mTORC2 but have yielded several conflicting hypotheses. More specifically, the Phosphoinositide 3-kinase (PI3K) pathway was shown to be involved in this process, but the identity of the kinase interacting with and regulating mTORC2 remains to be determined (Cybulski and Hall, Trends Biochem Sci 34:620-7, 2009). Method We performed a literature search and identified 5 published hypotheses describing mTORC2 regulation. Based on these hypotheses, we built logical models, not only for each single hypothesis but also for all combinations and possible mechanisms among them. Based on data provided by the original studies, a systematic analysis of all models was performed. Results We were able to find models that account for experimental observations from every original study, but do not require all 5 hypotheses to be implemented. Surprisingly, all hypotheses were in agreement with all tested data gathered from the different studies and PI3K was identified as an essential regulator of mTORC2. Conclusion The results and additional data suggest that more than one regulator is necessary to explain the behavior of mTORC2. Finally, this study proposes a new experiment to validate mTORC1 as second essential regulator

Global self-excited oscillations in a two-dimensional heated jet : a numerical simulation

The aim of this work was to develop a numerical methodology to gain insight in the low-density jet behaviour with a nonlinear approach. Numerical simulations are shown to differentiate convective and absolute instability regimes and to capture a self-excited global mode in an open flow : the 2D hot jet. The first part is devoted to numerical methodology and its validation on this unsteady problem which is known to be noise sensitive. The second part presents numerical results. They confirm theoretical and experimental results on the development of self-exited global oscillations of the jet column when the density ratio is lower than its critical value. The global mode and its associated Hopf bifurcation are identified

DNA copy number changes define spatial patterns of heterogeneity in colorectal cancer

Genetic heterogeneity between and within tumours is a major factor determining cancer progression and therapy response. Here we examined DNA sequence and DNA copy-number heterogeneity in colorectal cancer (CRC) by targeted high-depth sequencing of 100 most frequently altered genes. In 97 samples, with primary tumours and matched metastases from 27 patients, we observe inter-tumour concordance for coding mutations; in contrast, gene copy numbers are highly discordant between primary tumours and metastases as validated by fluorescent in situ hybridization. To further investigate intra-tumour heterogeneity, we dissected a single tumour into 68 spatially defined samples and sequenced them separately. We identify evenly distributed coding mutations in APC and TP53 in all tumour areas, yet highly variable gene copy numbers in numerous genes. 3D morpho-molecular reconstruction reveals two clusters with divergent copy number aberrations along the proximal–distal axis indicating that DNA copy number variations are a major source of tumour heterogeneity in CRC

Coopération scientifique sur la sécurité alimentaire, nutritionnelle et l'agriculture durable entre l'Afrique et l'Europe

Dans ce chapitre, les auteurs font valoir que la coopération scientifique sur la sécurité alimentaire, nutritionnelle et l'agriculture durable entre l'Afrique et l'Union européenne fait encore face à deux défis majeurs : d'une part, la mise en pratique de solutions innovantes est encore en retard; d'autre-part, cette coopération doit évoluer vers des partenariats de long terme, construits autour d'une gouvernance partagée. La sécurité alimentaire et nutritionnelle est soumise à des contraintes structurelles, socio-économiques, politiques et même environnementales qui, pour être levées, nécessitent d'importantes collaborations entre tous les acteurs de la société et dans divers secteurs d'activités, incluant la recherche et l'innovation. Les asymétries dans la coopération euro-africaine s'atténuent, mais elles doivent disparaître pour mieux contribuer à la réalisation des objectifs de la sécurité alimentaire et nutritionnelle en Afrique subsaharienne. Les auteurs soulignent le rôle stratégique que les pays africains peuvent jouer en diversifiant leur coopération notamment avec les pays européens moins présents dans les relations entre les deux continents et en promouvant des modèles de coopération innovants pour élaborer et développer des solutions concrètes aux problèmes alimentaires, nutritionnels et environnementaux sur la scène mondiale

Uncoupling of EGFR–RAS signaling and nuclear localization of YBX1 in colorectal cancer

The transcription factor YBX1 can act as a mediator of signals transmitted via the EGFR–RAS–MAPK axis. YBX1 expression has been associated with tumor progression and prognosis in multiple types of cancer. Immunohistochemical studies have revealed dependency between YBX1 expression and individual EGFR family members. We analyzed YBX1 and EGFR family proteins in a colorectal cancer (CRC) cohort and provide functional analyses of YBX1 in the context of EGFR–RAS–MAPK signaling. Immunohistochemistry for YBX1 and EGFR family receptors with two antibodies for YBX1 and EGFR were performed and related to clinicopathological data. We employed Caco2 cells expressing an inducible KRASV12 gene to determine effects on localization and levels of YBX1. Mouse xenografts of Caco2-KRASV12 cells were used to determine YBX1 dynamics in a tissue context. The two different antibodies against YBX1 showed discordant immunohistochemical stainings in cell culture and clinical specimens. Expression of YBX1 and EGFR family members were not correlated in CRC. Analysis of Caco2 xenografts displayed again heterogeneity of YBX1 staining with both antibodies. Our results suggest that YBX1 is controlled via complex regulatory mechanisms involving tumor stroma interaction and signal transduction processes. Our study highlights that YBX1 antibodies have different specificities, advocating their use in a combined manner

Funding of large-scale partnerships for co-owner-ship; what works and what doesn't

Policy brief, PROIntensAfrica201

Mutations of KRAS, NRAS, BRAF, EGFR, and PIK3CA genes in urachal carcinoma - occurence and prognostic significance

PURPOSE: Targeted therapy represents an attractive alternative for rare tumors such as urachal carcinoma (UrC). The aim of this study was to assess the mutations of the most commonly affected 5 genes in the targetable EGFR-pathway in UrC and comapre their frequencies to those of found in urothelial and colorectal cancer. MATERIALS AND METHODS: Mutational hot-spots of selected genes were tested in 22 UrC samples by pyrosequencing. Mutational patterns were compared to those published for colorectal and urothelial cancers. Furthermore, we sought correlations between mutations and clinicopathological and follow-up data. RESULTS: We found 11 mutations in 10 of 22 (45%) patients. The most frequently mutated gene was KRAS (27%) followed by BRAF (18%) and NRAS (5%), while no mutations were detected in the EGFR and PIK3CA genes. No correlation was found between the mutation status and clinicopathological parameters (Sheldon/Mayo stage, tumor grade, metastases). Furthermore, none of the mutations correlated with progression-free or overall survival. CONCLUSIONS: The mutation pattern of UrC is more similar to colorectal than to urothelial cancer. However, the mutation characteristics of UrC seems to be unique suggesting that clinical decision-making for UrC cannot be simply adopted from urothelial or colorectal carcinoma. The high occurence of EGFR-pathway mutations warrants the testing for KRAS and BRAF mutations when considering anti-EGFR therapy in UrC

Zielgene der RAS-Onkoprotein-abhängigen Signaltransduktion

Die Entstehung und Progression maligner Tumoren ist ein mehrstufiger Prozeß, der auf einer Vielzahl genetischer Alterationen beruht. Essentielle Schritte sind die Aktivierung von Proto-Onkogenen und die Inaktivierung von Tumor-Suppressorgenen. Infolge dessen können die Zellen unabhängig von externen Wachstumssignalen ungebremst proliferieren, die Apoptose wird gehemmt, die Angiogenese wird aktiviert, und es kommt schließlich zur Metastasierung. Zu den bekanntesten Proto-Onkogenen, die in humanen Tumoren aktiviert werden, gehören die RAS Gene. Sie sind in einer Vielzahl von Tumoren mutiert und führen zu einer Stimulation der Proliferation. Um den Einfluß aktivierter RAS Onkogene auf die Regulation der Genexpression zu untersuchen wurden Genexpressionsprofile in Zellkultur-Modellen und humanen Tumoren erstellt. In einem Fibroblasten- und einem Epithelzell-basierten System konnten mehrere hundert, RAS-abhängig differenziell exprimierte Genen identifiziert werden. Aufgrund der bekannten Funktionen ihrer Genprodukte spielen sie eine wichtige Rolle im Verlust der Zellzyklus-Kontrolle, der Kontrolle der Signalübertragung, in der Angiogenese-Induktion sowie in der Invasion und damit Metastasierung. Die Zusammenhänge zwischen der Aktivierung bestimmter Signalkaskaden wie z.B. Raf-Mek-Erk oder PI-3K und der Expression von definierten Genmustern wurden hergestellt. Weiterhin konnte mit Hilfe von Microarray Analysen eine Vielzahl potentieller Tumormarker und Zielgene für therapeutische Intervention im Ovarialkarzinom identifiziert werden. Die Rolle der KlasseII Tumorsuppressor Gene Caveolin-1 und H-REV107-1 in humanen Ovarialkarzinomen wurde detailliert untersucht und ihre Rolle in der Regulation des Zellüberlebens nachgewiesen. Caveolin-1, ein negativer Regulator der RAS-abhängigen Signalübertragung, wird in über 80% der untersuchten humanen Ovarialkarzinome gehemmt. Hierbei spielen epigenetische Mechanismen eine Rolle, die jedoch nicht Caveolin-1 selbst, sondern einen unbekannten Regulator des Caveolin-1 Gens betreffen. Das H-REV107-1 Gen, ein Wachstumsregulator mit unbekannter Funktion wird in ca. 50% der untersuchten Ovarialkarzinome nicht mehr exprimiert. Ähnlich wie bei Caveolin-1, führt eine gezielte Expression des Gens in Tumorzellen zur Apoptose. Die Suche nach Interaktionspartnern des H-REV107-1 Gens führte zur Identifizierung der ubiquitär exprimierten Phosphatase2A (PP2A). Die Bindung zwischen H-REV107-1 und PP2A wurde weiter charakterisiert und ihre Rolle in der H-REV107-1 vermittelten Apoptose analysiert.Development and progression of human tumours is a multistep process depending on numerous genetic alterations. Essentiell steps herein are the mutational activation of oncogenes and the inactivation of tumour suppressor genes. As a result of these alterations, the cells acquire the potential of unlimited growth independent of external growth factor signals, apoptosis is diminished, angiogenesis is stimulated and finally metastasis can occur. Among the best known proto-oncogenes, mutated in a number of human tumours, are the RAS genes. To investigate the role of RAS oncogenes in transformation-related transcriptional alterations, expressionsprofiling was performed from cell culture models and human tumours. Several hundred genes were identified to be de-regulated in a RAS-dependent manner in a fibroblast and an epithelial cell-based model. The protein products encoded by these genes play important roles in the loss of cell cycle control, control of signal transduction, angiogenesis induction as well as invasion and metastasis. Groups of de-regulated genes could be assigned to distinct signaling pathways such as the Raf-Mek-Erk or the PI-3 kinase dependent pathways. In addition, a number of potential tumour markers and potential target structures for therapeutic intervention were identified in ovarian carcinomas with the help of microarray studies. The role of the class II tumor suppressor genes Caveolin-1 and H-REV107-1 in human ovarian carcinomas was further investigated and their role in the regulation of cell survival was demonstrated. Caveolin-1, a negative regulator of RAS-dependent signal transduction, is supressed in more than 80% of the ovarian carcinomas analysed. This suppression is mediated by epigenetic mechanisms which due not target Caveolin-1 itself but an unknown regulator of the Caveolin-1 gene. The H-Rev107-1 gene, a growth regulator with unknown function, is no longer expressed in nearly 50% of the ovarian carcinomas analysed. Similar to Caveolin-1, also re-expression of H-REV107-1 results in apoptosis in the tumour cells. The search for proteins interacting with H-REV107-1 led to the identification of the ubiquitously expressed phosphatase 2A (PP2A). The interaction between H-REV107-1 and PP2A was further characterised and its role in the H-REV107-1 mediated apoptosis investigated

Evaluating Uncertainty in Signaling Networks Using Logical Modeling

Systems biology studies the structure and dynamics of biological systems using mathematical approaches. Bottom-up approaches create models from prior knowledge but usually cannot cope with uncertainty, whereas top-down approaches infer models directly from data using statistical methods but mostly neglect valuable known information from former studies. Here, we want to present a workflow that includes prior knowledge while allowing for uncertainty in the modeling process. We build not one but all possible models that arise from the uncertainty using logical modeling and subsequently filter for those models in agreement with data in a top-down manner. This approach enables us to investigate new and more complex biological research questions, however, the encoding in such a framework is often not obvious and thus not easily accessible for researcher from life sciences. To mitigate this problem, we formulate a pipeline with specific templates to address some research questions common in signaling network analysis. To illustrate the potential of this approach, we applied the pipeline to growth factor signaling processes in two renal cancer cell lines. These two cell lines originate from similar tissue, but surprisingly showed a very different behavior toward the cancer drug Sorafenib. Thus our aim was to explore differences between these cell lines regarding three sources of uncertainty in one analysis: possible targets of Sorafenib, crosstalk between involved pathways, and the effect of a mutation in mammalian target of Rapamycin (mTOR) in one of the cell lines. We were able to show that the model pools from the cell lines are disjoint, thus the discrepancies in behavior originate from differences in the cellular wiring. Also the mutation in mTOR is not affecting its activity in the pathway. The results on Sorafenib, while not fully clarifying the mechanisms involved, illustrate the potential of this analysis for generating new hypotheses.Peer Reviewe