Identification and characterization of genes involved in mouse embryonic stem cell differentiation into neurons

Mouse Embryonic Stem (ES) cells are pluripotent, as they have the ability to differentiate into all cell types of the embryo. Many protocols have been established to differentiate ES cells into the desired cell type in vitro. However, the identification of molecular machinery involved in ES cell biology will increase the knowledge of embryo development and will improve the use of ES cell for regenerative medicine. To identify genes involved in pluripotency and differentiation we developed a systematic approach based on the screening of a shRNA library. The results of the screening shed light on an uncharacterized gene, we named Dies1 (Differentiation of Embryonic Stem cells) given its effect on ES cell differentiation. Dies1 knock-down affected proper ES cell differentiation into neurons, astrocytes and cardiomyocytes. Dies1 knock-down cells, in fact, maintained the expression of undifferentiated markers such as Oct3/4, Nanog and Alkaline Phosphatase in conditions promoting differentiation. In addition, the growth of Dies1 knock-down cells was independent from LIF. Dies1 suppression exerted its effect on ES cell differentiation in vivo and in vitro, giving rise to teratoma smaller than controls when Dies1 knock-down cells are injected into nude mice. The in silico analysis of Dies1 sequence indicated that it possesses a trans-membrane domain and an Ig-like domain in the putative extra-cellular domain of the protein, like many membrane receptors. This analysis was confirmed by the staining of Dies1 on cell surface of mouse ES cell and blastocyst. The possible role for Dies1 as receptor was also supported by the dominant negative function of Dies1 extra-cellular domain, whose effect resembled Dies1 suppression. Thus, we investigated the involvement of Dies1 in the signaling pathways controlling ES cell pluripotency: LIF and BMP4. Dies1 knock-down did not affect the expression of undifferentiated markers such as Oct3/4, Nanog, Sox2 and Rex1 and of LIF down-stream targets such as Kruppel like factors in undifferentiated conditions. On the other hand, Inhibitory of Differentiation (Id) genes, which are BMP4 targets, were down-regulated in Dies1 knock-down cells. This down-regulation was dependent on a decreased response to BMP4, as shown by luciferase assays and occurred in a Smad dependent manner, as the amount of phosphorylated Smad 1/5/8 complex was decreased in Dies1 knock-down cells. Moreover, the involvement of Dies1 in BMP4 signaling was supported by the direct interaction between Dies1 and BMP4 in vitro. The down-regulation of BMP4 targets induced also the up-regulation of Nodal/Activin targets, such as Cripto and Lefty1/2. Such a regulation between Nodal/Activin and BMP4 was observed also in consequence of the suppression by RNA interference of the BMP4-type I receptor Alk3. This phenomenon was not surprising because Nodal/Activin and BMP4 belong to the TGF- superfamily and share the common mediator Smad4 which balances the transcriptional activity of Smad 2/3 (dependent on Nodal/Activin) and Smad 1/5/8 (dependent on BMPs). Thus, we hypothesized that the up-regulation of Nodal/Activin signaling was responsible for the effect of Dies1 suppression. To verify this hypothesis we treated Dies1 knock-down cells with the specific Nodal/Activin inhibitor SB-431542. The treatment with SB-431542 restored proper differentiation ability in the absence of LIF, leading to the rescue of Dies1 knock-down effect. Taken together our results suggest that the balance between BMP4 and Nodal/Activin exerts a crucial role in regulating ES cell pluripotency and differentiation. In summary, we have identified Dies1 as novel component of BMP4 signaling pathway required for proper ES cell differentiation

Introduction

Introduction to Building Bridges: Tools for Open Educational Resources (OER) Advocacy and Collaboration at TCUS Libraries , an online workshop held April 13th, 2021 1:00 P.M. - 4:00 P.M. Eastern Time

Epigenetic Regulation of Cell-Fate Changes That Determine Adult Liver Regeneration After Injury

The adult liver has excellent regenerative potential following injury. In contrast to other organs of the body that have high cellular turnover during homeostasis (e.g., intestine, stomach, and skin), the adult liver is a slowly self-renewing organ and does not contain a defined stem-cell compartment that maintains homeostasis. However, tissue damage induces significant proliferation across the liver and can trigger cell-fate changes, such as trans-differentiation and de-differentiation into liver progenitors, which contribute to efficient tissue regeneration and restoration of liver functions. Epigenetic mechanisms have been shown to regulate cell-fate decisions in both embryonic and adult tissues in response to environmental cues. Underlying their relevance in liver biology, expression levels and epigenetic activity of chromatin modifiers are often altered in chronic liver disease and liver cancer. In this review, I examine the role of several chromatin modifiers in the regulation of cell-fate changes that determine efficient adult liver epithelial regeneration in response to tissue injury in mouse models. Specifically, I focus on epigenetic mechanisms such as chromatin remodelling, DNA methylation and hydroxymethylation, and histone methylation and deacetylation. Finally, I address how altered epigenetic mechanisms and the interplay between epigenetics and metabolism may contribute to the initiation and progression of liver disease and cancer

The influence of tissue spatial geometry and functional organisation on liver regeneration

The adult liver exerts crucial functions, including nutrient metabolism and storage, bile production and drug detoxification. These complex functions expose the liver to constant damage induced by toxins, metabolic intermediates and oxidative stress. However, the adult liver exhibits an exceptional regenerative potential, which allows fast and efficient restoration of tissue architecture and function both after tissue resection and toxic damage. To accomplish its vital role, the liver shows a peculiar tissue architecture into functional units, which follow the gradient of oxygen and nutrients within the parenchyma. Much less is known about the influence of tissue spatial geometry and functional organisation on adult liver regeneration. Here I examine the experimental evidence in mouse models showing that the spatial organisation of the epithelial and mesenchymal compartments plays a key role in liver regeneration and favours the establishment of regenerative adult liver progenitors following liver injury. I also discuss the advantages and limitations of human and mouse 3D hepatic organoid systems, which recapitulate key aspects of liver function and architecture, as models of liver regeneration and disease. Finally, I analyse the role of the YAP/TAZ transcriptional co-activators as a central hub sensing the extra-cellular matrix (ECM), metabolic and epigenetic remodelling that regulate liver regeneration and promote liver disease, such as fibrosis, chronic liver disease and liver cancer. Together, the findings summarised here demonstrate that local physical and functional cellular interactions determined by the liver peculiar spatial geometry, play a crucial role in liver regeneration, and that their alterations have important implications for human liver disease

Eucalyptus wood and cheese whey valorization for biofuels production

In this work, two raw materials (Eucalyptus wood and cheese whey) were used for ethanol production. Eucalyptus wood was hydrothermally pretreated at 233 ºC in order to increase the enzymatic saccharification of cellulose. Pretreated Eucalyptus wood mixed or not with cheese whey were used as substrates for ethanol production by simultaneous saccharification and fermentation (SSF) using two Saccharomyces cerevisiae strains (industrial Ethanol Red® and laboratory CEN.PK1137D). The use of cheese whey mixed with Eucalyptus wood increased 1.3 and 1.5-fold the ethanol concentration in comparison with Eucalyptus without cheese whey using S. cerevisiae Ethanol Red® and CEN.PK113 7D strains, respectively. Higher ethanol concentration was obtained by Ethanol Red® than ethanol produced by CEN.PK113-7D with cheese whey supplementation (93 g/L and 65 g/L corresponding to 94 % and 66 % of ethanol yield, respectively). Results obtained in this work showed an interesting strategy for the valorization of two raw materials in order to produce high concentrations of ethanol.Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2013 unit and COMPETE 2020 (POCI -01-0145-FEDER-006684) and BioTecNorte operation (NORTE-01- 0145-FEDER -000004) funded by European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte.info:eu-repo/semantics/publishedVersio

Cellular plasticity in the adult liver and stomach.

Adult tissues maintain function and architecture through robust homeostatic mechanisms mediated by self-renewing cells capable of generating all resident cell types. However, severe injury can challenge the regeneration potential of such a stem/progenitor compartment. Indeed, upon injury adult tissues can exhibit massive cellular plasticity in order to achieve proper tissue regeneration, circumventing an impaired stem/progenitor compartment. Several examples of such plasticity have been reported in both rapidly and slowly self-renewing organs and follow conserved mechanisms. Upon loss of the cellular compartment responsible for maintaining homeostasis, quiescent or slowly proliferating stem/progenitor cells can acquire high proliferation potential and turn into active stem cells, or, alternatively, mature cells can de-differentiate into stem-like cells or re-enter the cell cycle to compensate for the tissue loss. This extensive cellular plasticity acts as a key mechanism to respond to multiple stimuli in a context-dependent manner, enabling tissue regeneration in a robust fashion. In this review cellular plasticity in the adult liver and stomach will be examined, highlighting the diverse cell populations capable of repairing the damaged tissue.MH is a Wellcome Trust Sir Henry Dale Fellow and is jointly funded by the Wellcome Trust and the Royal Society (104151/Z/14/Z). MM is an MRC PhD fellow (PMAG/440).This is the author accepted manuscript. The final version is available from Wiley via http://dx.doi.org/10.1113/JP27176

Environmentally sustainable processes for biomass conversion into biofuels and value-added compounds: integrated and intensified approach within a biorefinery concept

Book of Abstracts of CEB Annual Meeting 2017[Excerpt] Lignocellulosic biomass conversion into biofuels is considered a promising alternative to replace fossil fuels, being one of investment priorities of European Union to attain a sustainable growth within Horizon 2020. Nevertheless, lignocellulosic biofuels are not widely implemented on large-scale due to the high initial investment and operational costs. The scientific research carried out has been focused in the development of biomass processing technology for bioethanol production making use of environmentally-friendly pre-treatments and molecular biotechnology tools (metabolic, genetic and physiological engineering) for yeast development. [...]info:eu-repo/semantics/publishedVersio

Searching Data: A Review of Observational Data Retrieval Practices in Selected Disciplines

A cross-disciplinary examination of the user behaviours involved in seeking and evaluating data is surprisingly absent from the research data discussion. This review explores the data retrieval literature to identify commonalities in how users search for and evaluate observational research data. Two analytical frameworks rooted in information retrieval and science technology studies are used to identify key similarities in practices as a first step toward developing a model describing data retrieval