Separate functions of BTZ during post-­transcriptional gene regulation

In metazoans, the exon junction complex (EJC) is a central component of spliced messenger ribonucleoprotein particles (mRNPs). EJCs are assembled by the spliceosome and deposited upstream of exon-exon boundaries in the nucleus. The heterotetrameric core of the EJC is composed of the proteins eIF4A3 (DDX48), MAGOH, RBM8 (Y14) and CASC3/MLN51/Barentsz (BTZ). EJCs contribute to different steps of post-transcriptional gene expression including splicing, translation and nonsense-mediated mRNA decay (NMD). BTZ is an important functional component and is involved in the stimulation of translation and nonsense-mediated mRNA decay. Here, I show that both the N-terminal and the SELOR domain of BTZ elicit NMD in a tethering assay. They activate NMD following two different pathways, BTZ-dependent and UPF2 dependent, which get reunited once UPF1 is activated. In contrast, the C-terminal region of BTZ does not seem to be involved in NMD. Instead, this region plays a role in a different process that leads to the polyadenylation of a reporter mRNA at an upstream, non-canonical polyadenylation site. Moreover, I show that binding of the SELOR domain to mRNA in vivo is EJC-dependent. In addition the SELOR domain in vivo interacts with several SR proteins for a subset of which an NMD-activating function is observed. These findings uncover novel EJC-dependent and -independent functions of BTZ during post-transcriptional gene expression regulation

Impact of space-time mesh adaptation on solute transport modeling in porous media

open4siWe implement a space-time grid adaptation procedure to efficiently improve the accuracy of numerical simulations of solute transport in porous media in the context of model parameter estimation. We focus on the Advection Dispersion Equation (ADE) for the interpretation of non-reactive transport experiments in laboratory-scale heterogeneous porous media. When compared to a numerical approximation based on a fixed space-time discretization, our approach is grounded on a joint automatic selection of the spatial grid and the time step to capture the main (space-time) system dynamics. Spatial mesh adaptation is driven by an anisotropic recovery-based error estimator which enables us to properly select the size, shape and orientation of the mesh elements. Adaptation of the time step is performed through an ad-hoc local reconstruction of the temporal derivative of the solution via a recovery-based approach. The impact of the proposed adaptation strategy on the ability to provide reliable estimates of the key parameters of an ADE model is assessed on the basis of experimental solute breakthrough data measured following tracer injection in a non-uniform porous system. Model calibration is performed in a Maximum Likelihood (ML) framework upon relying on the representation of the ADE solution through a generalized Polynomial Chaos Expansion (gPCE). Our results show that the proposed anisotropic space-time grid adaptation leads to ML parameter estimates and to model results of markedly improved quality when compared to classical inversion approaches based on a uniform space-time discretization.openEsfandiar, B; Porta, G; Perotto, S; Guadagnini, AESFANDIAR JAHROMI, Bahman; Porta, GIOVANNI MICHELE; Perotto, Simona; Guadagnini, Albert

Direct targets of Klf5 transcription factor contribute to the maintenance of mouse embryonic stem cell undifferentiated state

<p>Abstract</p> <p>Background</p> <p>A growing body of evidence has shown that Krüppel-like transcription factors play a crucial role in maintaining embryonic stem cell (ESC) pluripotency and in governing ESC fate decisions. Krüppel-like factor 5 (Klf5) appears to play a critical role in these processes, but detailed knowledge of the molecular mechanisms of this function is still not completely addressed.</p> <p>Results</p> <p>By combining genome-wide chromatin immunoprecipitation and microarray analysis, we have identified 161 putative primary targets of Klf5 in ESCs. We address three main points: (1) the relevance of the pathways governed by Klf5, demonstrating that suppression or constitutive expression of single Klf5 targets robustly affect the ESC undifferentiated phenotype; (2) the specificity of Klf5 compared to factors belonging to the same family, demonstrating that many Klf5 targets are not regulated by Klf2 and Klf4; and (3) the specificity of Klf5 function in ESCs, demonstrated by the significant differences between Klf5 targets in ESCs compared to adult cells, such as keratinocytes.</p> <p>Conclusions</p> <p>Taken together, these results, through the definition of a detailed list of Klf5 transcriptional targets in mouse ESCs, support the important and specific functional role of Klf5 in the maintenance of the undifferentiated ESC phenotype.</p> <p>See: <url>http://www.biomedcental.com/1741-7007/8/125</url></p

Studio dei processi di emocoagulazione mediante Quartz Crystal Microbalance

Lo studio ed il controllo dei processi di fibrinolisi e coagulazione del sangue rivestono un vasto interesse medico e clinico in particolar modo per quei pazienti con patologie cardiovascolari, portatori di protesi cardiache ed in tutti quei trattamenti nei quali il sangue entra a contatto con superfici artificiali. L'impiego di anticoagulanti, tipicamente utilizzati in questi casi per prevenire o minimizzare gli indesiderati ma inevitabili effetti di clothing richiede il frequente controllo di alcuni parametri ematici (ad es. tempo di coagulazione) pena l'instaurarsi di una serie di patologie che possono gravemente compromettere la salute del paziente. Il tempo di coagulazione del sangue viene tipicamente valutato dalla misura della variazione della sua viscosità su processi di coagulazione indotti mediante attivatori interfasali o enzimi. Quantunque esistano ben note procedure cliniche strumentali ad hoc, recentemente si sta affacciando l'approccio alternativo basato sui biosensori [1-3], nell'intento di produrre dispositivi avanzati in grado di fornire con tempestività e precisioni parametri ematoclinici anche in regime di self-analysis. Il presente lavoro di ricerca è proprio rivolto allo sviluppo ed utilizzo di un originale sensore basato su microbilancia a cristallo di quarzo, in grado di seguire le cinetiche di emocoagulazione mediante la deposizione del campione stesso sulla superficie del cristallo e la contestuale acquisizione dei relativi spettri di ammittanza. Il sistema messo a punto è in grado di seguire il processo di coagulazione di campioni di sangue del volume di pochi microlitri sia indotto da superficie sia tramite trombina. In plasma umano, la fase di coagulazione è evidenziata da una decisa diminuzione dei picchi di ammittanza con parallela diminuzione della frequenza di risonanza e del fattore Q del sistema cristallo di quarzo - campione di sangue, con cinetiche analoghe a quelle osservate con procedure strumentali standard. Contrariamente a quest'ultime, viceversa, il sensore sviluppato nel presente lavoro di ricerca offre anche la possibilità di evidenziare l'ulteriore processo conseguente la produzione di fibrina ossia la formazione del vero e proprio coagulo, e di fornire utili indicazione sul grado di rigidità dello stesso. Quest'ultimo aspetto appare poco esplorato nella relativa letteratura scientifica e quindi rappresenta un aspetto fortemente originale della presente ricerca. [1] Hansson, K.M. Vikinge, T.P. Ronby, M. Tengvall, P. Lundstrom, L. Johansen, K. Lindahl, T.L., Biosens. Bioelectron., 14, 671 (1999). [2] Puckett, L.G. Barrett, G. Kouzoudis, D. Grimes, C. Bachas, L.G., Biosens. Bioelectron., 18, 675 (2003). [3] Andersson, M. Andersson, J. Sellborn, A. Berglin, M. Nilsson, B. Elwing, H., Biosens. Bioelectron., 21, 79 (2005)

CASC3 promotes transcriptome-wide activation of nonsense-mediated decay by the exon junction complex

The exon junction complex (EJC) is an essential constituent and regulator of spliced messenger ribonucleoprotein particles (mRNPs) in metazoans. As a core component of the EJC, CASC3 was described to be pivotal for EJC-dependent nuclear and cytoplasmic processes. However, recent evidence suggests that CASC3 functions differently from other EJC core proteins. Here, we have established human CASC3 knockout cell lines to elucidate the cellular role of CASC3. In the knockout cells, overall EJC composition and EJC-dependent splicing are unchanged. A transcriptome-wide analysis reveals that hundreds of mRNA isoforms targeted by nonsense-mediated decay (NMD) are upregulated. Mechanistically, recruiting CASC3 to reporter mRNAs by direct tethering or via binding to the EJC stimulates mRNA decay and endonucleolytic cleavage at the termination codon. Building on existing EJC-NMD models, we propose that CASC3 equips the EJC with the persisting ability to communicate with the NMD machinery in the cytoplasm. Collectively, our results characterize CASC3 as a peripheral EJC protein that tailors the transcriptome by promoting the degradation of EJC-dependent NMD substrates

Data_Sheet_1_Substrate and fertigation management modulate microgreens production, quality and resource efficiency.pdf

Long-term space missions will require a self-sustaining food production system to meet the crew’s nutritional and health needs. For this purpose, plant-based food production systems with elevated resource efficiency are required, based on advanced agricultural technologies that produce phytonutrient-rich crops. In addition to the resource requirements for crop production on Earth, volume and time efficiency become essential factors to consider for food production in space. Microgreens represent a promising candidate for space farming as they have a high harvest index, short cultivation cycle, and high nutritional potential. However, the development of specific technical protocols for growing microgreens in space is essential since different agronomic inputs, such as substrates and fertigation, can modulate productivity, quality and resource efficiency of microgreens cultivation. The current work examines the effects of different substrates (coconut fiber and cellulose sponge) and nutrient solution (NS) management strategies (quarter strength Hoagland and half strength Hoagland/osmotic water) on the production of two species of microgreens [Raphanus sativus cv. Saxa 2 (Radish); Brassica oleracea var. capitata f. sabauda cv. Vertus (Savoy cabbage)]. The appraisal focused on (i) biomass production and quality, and (ii) sizing of space facilities devoted to the production of phytonutrients required for the astronauts’ wellbeing. In our study, the interaction among species, substrate and NS significantly affected the accumulation of fructose, sucrose, total soluble non-structural carbohydrates and nitrate as well as the daily production of total ascorbic acid and, in turn, the required microgreens serving to supply its adequate daily intake. Species-substrate interaction effects on fresh yield, dry yield, dry matter, anthocyanins, TPC, β-carotene and sulfate content as well as the cultivation surface required to produce the adequate daily intake of ascorbic acid (AscA) were assessed. Substrate-NS interaction modulated the anthocyanins, violaxanthin and sulfate contents independently of species. On the other hand, single factor effects were identified with respect to the accumulation of lutein, chlorophylls, glucose, and starch. Therefore, the management of microgreens cultivation in terms of NS and substrate is an effective tool to meet the phytochemical requirements of the crew.</p

Liver Activation of Hepatocellular Nuclear Factor-4α by Small Activating RNA Rescues Dyslipidemia and Improves Metabolic Profile

Non-alcoholic fatty liver disease (NAFLD) culminates in insulin resistance and metabolic syndrome. Because there are no approved pharmacological treatment agents for non-alcoholic steatohepatitis (NASH) and NAFLD, different signaling pathways are under investigation for drug development with the focus on metabolic pathways. Hepatocyte nuclear factor 4-alpha (HNF4A) is at the center of a complex transcriptional network where its disruption is directly linked to glucose and lipid metabolism. Resetting HNF4A expression in NAFLD is therefore crucial for re-establishing normal liver function. Here, small activating RNA (saRNA) specific for upregulating HNF4A was injected into rats fed a high-fat diet for 16 weeks. Intravenous delivery was carried out using 5-(G5)-triethanolamine-core polyamidoamine (PAMAM) dendrimers. We observed a significant reduction in liver triglyceride, increased high-density lipoprotein/low-density lipoprotein (HDL/LDL) ratio, and decreased white adipose tissue/body weight ratio, all parameters to suggest that HNF4A-saRNA treatment induced a favorable metabolic profile. Proteomic analysis showed significant regulation of genes involved in sphingolipid metabolism, fatty acid β-oxidation, ketogenesis, detoxification of reactive oxygen species, and lipid transport. We demonstrate that HNF4A activation by oligonucleotide therapy may represent a novel single agent for the treatment of NAFLD and insulin resistance