Genome-Wide Analysis of Smad7-Mediated Transcription in Mouse Embryonic Stem Cells.

Smad7 has been identified as a negative regulator of the transforming growth factor TGF-ß pathway by direct interaction with the TGF-ß type I receptor (TßR-I). Although Smad7 has also been shown to play TGF-ß unrelated functions in the cytoplasm and in the nucleus, a comprehensive analysis of its nuclear function has not yet been performed. Here, we show that in ESCs Smad7 is mainly nuclear and acts as a general transcription factor regulating several genes unrelated to the TGF-ß pathway. Loss of Smad7 results in the downregulation of several key stemness master regulators, including Pou5f1 and Zfp42, and in the upregulation of developmental genes, with consequent loss of the stem phenotype. Integrative analysis of genome-wide mapping data for Smad7 and ESC self-renewal and pluripotency transcriptional regulators revealed that Smad7 co-occupies promoters of highly expressed key stemness regulators genes, by binding to a specific consensus response element NCGGAAMM. Altogether, our data establishes Smad7 as a new, integral component of the regulatory circuitry that controls ESC identity.

A Comprehensive Bioinformatics Analysis of UBE2C in Cancers.

Ubiquitination is one of the main post-translational modification of proteins. It plays key roles in a broad range of cellular functions, including protein degradation, protein interactions, and subcellular location. In the ubiquitination system, different proteins are involved and their dysregulation can lead to various human diseases, including cancers. By using data available from the Cancer Genome Atlas (TCGA) and the Genotype-Tissue Expression (GTEx) databases, we here show that the ubiquitin conjugating enzyme, E2C (UBE2C), is overexpressed in all 27 cancers we investigated. UBE2C expression is significantly higher in late-stage tumors, which might indicate its involvement in tumor progression and invasion. This study also revealed that patients with higher UBE2C levels showed a shorter overall survival (OS) time and worse OS prognosis. Moreover, our data show that UBE2C higher-expression leads to worse disease-free survival prognosis (DFS), indicating that UBE2C overexpression correlates with poor clinical outcomes. We also identified genes with positive correlations with UBE2C in several cancers. We found a number of poorly studied genes (family with sequence similarity 72-member D, FAM72D; meiotic nuclear divisions 1, MND1; mitochondrial fission regulator 2, MTFR2; and POC1 centriolar protein A, POC1A) whose expression correlates with UBE2C. These genes might be considered as new targets for cancers therapies since they showed overexpression in several cancers and correlate with worse OS prognosis.

DNMT3B supports meso-endoderm differentiation from mouse embryonic stem cells.

The correct establishment of DNA methylation patterns during mouse early development is essential for cell fate specification. However, the molecular targets as well as the mechanisms that determine the specificity of the de novo methylation machinery during differentiation are not completely elucidated. Here we show that the DNMT3B-dependent DNA methylation of key developmental regulatory regions at epiblast-like cells (EpiLCs) provides an epigenetic priming that ensures flawless commitment at later stages. Using in vitro stem cell differentiation and loss of function experiments combined with high-throughput genome-wide bisulfite-, bulk-, and single cell RNA-sequencing we dissected the specific role of DNMT3B in cell fate. We identify DNMT3B-dependent regulatory elements on the genome which, in Dnmt3b knockout (3BKO), impair the differentiation into meso-endodermal (ME) progenitors and redirect EpiLCs towards the neuro-ectodermal lineages. Moreover, ectopic expression of DNMT3B in 3BKO re-establishes the DNA methylation of the master regulator Sox2 super-enhancer, downmodulates its expression, and restores the expression of ME markers. Taken together, our data reveal that DNMT3B-dependent methylation at the epiblast stage is essential for the priming of the meso-endodermal lineages and provide functional characterization of the de novo DNMTs during EpiLCs lineage determination.

Tissue memory relies on stem cell priming in distal undamaged areas.

Epithelial cells that participated in wound repair elicit a more efficient response to future injuries, which is believed to be locally restricted. Here we show that cell adaptation resulting from a localized tissue damage has a wide spatial impact at a scale not previously appreciated. We demonstrate that a specific stem cell population, distant from the original injury, originates long-lasting wound memory progenitors residing in their own niche. Notably, these distal memory cells have not taken part in the first healing but become intrinsically pre-activated through priming. This cell state, maintained at the chromatin and transcriptional level, leads to an enhanced wound repair that is partially recapitulated through epigenetic perturbation. Importantly wound memory has long-term harmful consequences, exacerbating tumourigenesis. Overall, we show that sub-organ-scale adaptation to injury relies on spatially organized memory-dedicated progenitors, characterized by an actionable cell state that establishes an epigenetic field cancerization and predisposes to tumour onset.

IFNγ-Stat1 axis drives aging-associated loss of intestinal tissue homeostasis and regeneration.

The influence of aging on intestinal stem cells and their niche can explain underlying causes for perturbation in their function observed during aging. Molecular mechanisms for such a decrease in the functionality of intestinal stem cells during aging remain largely undetermined. Using transcriptome-wide approaches, our study demonstrates that aging intestinal stem cells strongly upregulate antigen presenting pathway genes and over-express secretory lineage marker genes resulting in lineage skewed differentiation into the secretory lineage and strong upregulation of MHC class II antigens in the aged intestinal epithelium. Mechanistically, we identified an increase in proinflammatory cells in the lamina propria as the main source of elevated interferon gamma (IFNγ) in the aged intestine, that leads to the induction of Stat1 activity in intestinal stem cells thus priming the aberrant differentiation and elevated antigen presentation in epithelial cells. Of note, systemic inhibition of IFNγ-signaling completely reverses these aging phenotypes and reinstalls regenerative capacity of the aged intestinal epithelium.

Interplay between the endocannabinoid system and GnRH-I in the forebrain of the anuran amphibian Rana esculenta.

The morphofunctional relationship between the endocannabinoid system and GnRH activity in the regulation of reproduction has poorly been investigated in vertebrates. Due to the anatomical features of lower vertebrate brain, in the present paper, we chose the frog Rana esculenta (anuran amphibian) as a suitable model to better investigate such aspects of the reproductive physiology. By using double-labeling immunofluorescence aided with a laser-scanning confocal microscope, we found a subpopulation of the frog hypothalamic GnRH neurons endowed with CB1 cannabinoid receptors. By means of semiquantitative RT-PCR assay, we have shown that, during the annual sexual cycle, GnRH-I mRNA (formerly known as mammalian GnRH) and CB1 mRNA have opposite expression profiles in the brain. In particular, this occurs in telencephalon and diencephalon, the areas mainly involved in GnRH release and control of the reproduction. Furthermore, we found that the endocannabinoid anandamide is able to inhibit GnRH-I mRNA synthesis; buserelin (a GnRH agonist), in turn, inhibits the synthesis of GnRH-I mRNA and induces an increase of CB1 transcription. Our observations point out the occurrence of a morphofunctional anatomical basis to explain a reciprocal relationship between the endocannabinoid system and GnRH neuronal activity.

Enzymatic changes in mitochondria-rich cells of Xenopus laevis skin epithelium are induced by ionic acclimation.

Morphological, biochemical and histochemical components of mitochondria-rich (MR) cells of skin epithelium of Xenopus laevis (Daudin) were investigated after acclimation in distilled water (DW) and mild solutions (50 mmol/l) of either NaCl or KCl for over 10 days. The animals readily acclimated to NaCl, but approximately 50% of the animals died in the KCl solution. Electrophysiological measurements confirmed the poor transport properties of skin in all conditions. Silver staining and exposure to methylene blue (MB) have shown that two types of MR cells can be distinguished, especially after KCl acclimation. Immunohistochemistry with the use of anti-band 3 polyclonal and anti H+-ATPase monoclonal antibodies demonstrated that band 3 and H+-ATPase enzymes were localized in MR cells in all conditions. H+-ATPase was greatly reduced during NaCl acclimation as verified with SDS gel electrophoresis. Intensity of the immunohistochemical staining differed between the various conditions of acclimation. Histochemical localization of carbonic anhydrase and alkaline phosphatase activities was more intense during NaCl acclimation. Morphological changes were also observed between the various acclimation conditions. The present findings substantiate the existence of at least two forms of MR cells in Xenopus skin epithelium but their functional significance remains to be established.