Explore the vast range of titles available.

miRWalk is an open-source platform providing an intuitive interface that generates predicted and validated miRNA-binding sites of known genes of human, mouse, rat, dog and cow. The core of miRWalk is the miRNA target site prediction with the random-forest-based approach software TarPmiR searching the complete transcript sequence including the 5'-UTR, CDS and 3'-UTR. Moreover, it integrates results other databases with predicted and validated miRNA-target interactions. The focus is set on a modular design and extensibility as well as a fast update cycle. The database is available using Python, MySQL and HTML/Javascript Database URL: http://mirwalk.umm.uni-heidelberg.de.

Tumors have aberrant proteomes that often do not match their corresponding transcriptome profiles. One possible cause of this discrepancy is the existence of aberrant RNA modification landscapes in the so-called epitranscriptome. Here, we report that human glioma cells undergo DNA methylation-associated epigenetic silencing of NSUN5, a candidate RNA methyltransferase for 5-methylcytosine. In this setting, NSUN5 exhibits tumor-suppressor characteristics in vivo glioma models. We also found that NSUN5 loss generates an unmethylated status at the C3782 position of 28S rRNA that drives an overall depletion of protein synthesis, and leads to the emergence of an adaptive translational program for survival under conditions of cellular stress. Interestingly, NSUN5 epigenetic inactivation also renders these gliomas sensitive to bioactivatable substrates of the stress-related enzyme NQO1. Most importantly, NSUN5 epigenetic inactivation is a hallmark of glioma patients with long-term survival for this otherwise devastating disease.

The nucleotide analogue azacitidine (AZA) is currently the best treatment option for patients with high-risk myelodysplastic syndromes (MDS). However, only half of treated patients respond and of these almost all eventually relapse. New treatment options are urgently needed to improve the clinical management of these patients. Here, we perform a loss-of-function shRNA screen and identify the histone acetyl transferase and transcriptional co-activator, CREB binding protein (CBP), as a major regulator of AZA sensitivity. Compounds inhibiting the activity of CBP and the closely related p300 synergistically reduce viability of MDS-derived AML cell lines when combined with AZA. Importantly, this effect is specific for the RNA-dependent functions of AZA and not observed with the related compound decitabine that is only incorporated into DNA. The identification of immediate target genes leads us to the unexpected finding that the effect of CBP/p300 inhibition is mediated by globally down regulating protein synthesis. Azacitidine (AZA) treatment is used for patients with myelodysplasias that cannot undergo bone marrow transplantation; however, AZA treatment is only partially effective. Here the authors show synergy of AZA with compounds inhibiting the chromatin regulators CBP and p300, which is mediated by the RNA-dependent functions of AZA affecting protein translation.

The oncogene yes-associated protein (YAP) is a key modifier of liver homeostasis and regulates mitosis in hepatocytes as well as in malignantly transformed cells. However, the question of how YAP supports cell proliferation in hepatocellular carcinoma (HCC) is not well understood. Here we identified U2AF momology motif kinase 1 (UHMK1) as a direct transcriptional target of YAP and the transcription factor forkhead box M1 (FOXM1), which supports HCC cell proliferation but not migration. Indeed, UHMK1 stimulates the expression of genes that are specific for cell cycle regulation and which are known downstream effectors of YAP. By using BioID labeling and mass spectrometry, the dimerization partner, RB-like, E2F and multi-vulval class B (DREAM) complex constituent MYB proto-oncogene like 2 (MYBL2, B-MYB) was identified as a direct UHMK1 interaction partner. Like YAP, UHMK1 stimulates nuclear enrichment of MYBL2, which is associated HCC cell proliferation and the expression of the cell cycle regulators CCNB1, CCNB2, KIF20A, and MAD2L1. The association between YAP, UHMK1, MYBL2, and proliferation was confirmed in YAP S127A -transgenic mice and human HCC tissues. In summary, we provide a model by which YAP supports cell proliferation through the induction of important cell cycle regulators in a UHMK1- and MYBL2-dependent manner.

Porcine intestinal mucosa hydrolysates (PIMHs) are by-products of heparin production obtained through a specific enzymatic hydrolysis process, which can theoretically generate bioactive peptides (BAPs). This study aimed to identify, characterize, and quantify BAPs in two Palbio products manufactured by Bioiberica S.A.U. (Palafolls, Spain), which are PIMH protein sources used for animal feed: Palbio® HP (PHP) and Palbio® 62 SP® (P62). Using mass spectrometry (MS)-based peptidomics, we analyzed three samples from each product, fractionated based on molecular weight (<3 kDa, 3 to 10 kDa, and >10 kDa). The <3 kDa fraction was analyzed directly, while the other two fractions were enzymatically digested before MS analysis. The workflow identified 961 peptides in PHP and 1134 in P62. Subsequent bioinformatic analysis using public databases (APD2, StraPep, AHTPDB, and BIOPEP-UWM) led to the identification of six significant BAPs in both PHP and P62, with respective quantified amounts (pg peptide/μg sample): DAVEDLESVGK (0.1626, 0.1939), EGIPPDQQRLIFAGK (0.2637, 0.1852), TITLEVEPSDTIENVK (0.3594, 0.4327), TNVPRASVPDGFLS (1.4596, 0.1898), TNVPRASVPDGFLSEL (8.0500, 0.9224), and VHVVPDQLMAF (0.0310, 0.0054). The first three BAPs are related to antimicrobial activity, while the latter three are associated with cytokine/growth factor-like, antioxidant, and immunomodulatory activities. These bioactivities align with previously reported in vivo benefits observed in animal nutrition using Palbio products. Our findings demonstrate that PHP and P62 are valuable sources of BAPs, supporting their potential role in improving animal health and performance.

NOTCH receptor signaling plays a pivotal role in liver homeostasis and hepatocarcinogenesis. However, the role of NOTCH pathway mutations and the NOTCH target gene HES5 in liver tumorigenesis are poorly understood. Here we performed whole-exome sequencing of 54 human HCC specimens and compared the prevalence of NOTCH pathway component mutations with the TCGA-LIHC cohort (N = 364). In addition, we functionally characterized the NOTCH target HES5 and the patient-derived HES5-R31G mutation in vitro and in an orthotopic mouse model applying different oncogenic backgrounds, to dissect the role of HES5 in different tumor subgroups in vivo. We identified nonsynonymous mutations in 14 immediate NOTCH pathway genes affecting 24.1% and 16.8% of HCC patients in the two independent cohorts, respectively. Among these, the HES5-R31G mutation was predicted in silico to have high biological relevance. Functional analyses in cell culture showed that HES5 reduced cell migration and clonogenicity. Further analyses revealed that the patient-derived HES5-R31G mutant protein was non-functional due to loss of DNA binding and greatly reduced nuclear localization. Furthermore, HES5 exhibited a negative feedback loop by directly inhibiting the NOTCH target HES1 and downregulated the pro-proliferative MYC targets ODC1 and LDHA. Interestingly, HES5 inhibited MYC-dependent hepatocarcinogenesis, whereas it promoted AKT-dependent liver tumor formation and stem cell features in a murine model. Thus, NOTCH pathway component mutations are commonly observed in HCC. Furthermore, the NOTCH target gene HES5 has both pro- and anti-tumorigenic functions in liver cancer proposing a driver gene dependency and it promotes tumorigenesis with its interaction partner AKT.

Within the bone marrow microenvironment, endothelial cells (EC) exert important functions. Arterial EC support hematopoiesis while H-type capillaries induce bone formation. Here, we show that BM sinusoidal EC (BM-SEC) actively control erythropoiesis. Mice with stabilized β-catenin in BM-SEC ( Ctnnb1 OE-SEC ) generated by using a BM-SEC-restricted Cre mouse line ( Stab2-iCreF3 ) develop fatal anemia. While activation of Wnt-signaling in BM-SEC causes an increase in erythroblast subsets (PII–PIV), mature erythroid cells (PV) are reduced indicating impairment of terminal erythroid differentiation/reticulocyte maturation. Transplantation of Ctnnb1 OE-SEC hematopoietic stem cells into wildtype recipients confirms lethal anemia to be caused by cell-extrinsic, endothelial-mediated effects. Ctnnb1 OE-SEC BM-SEC reveal aberrant sinusoidal differentiation with altered EC gene expression and perisinusoidal ECM deposition and angiocrine dysregulation with de novo endothelial expression of FGF23 and DKK2, elevated in anemia and involved in vascular stabilization, respectively. Our study demonstrates that BM-SEC play an important role in the bone marrow microenvironment in health and disease. Niche crosstalk with Haematopoietic cells underlies normal haematopoiesis and myeloid disorders. Here the authors report a Stabilin2-Cre driver mouse with Cre-activity restricted to bone marrow sinusoidal endothelial cells, and that Stabilin2-Cre driven overactivation of b-catenin leads to erythroid differentiation defects and anaemia.

Melanoma immunotherapy is still not satisfactory due to immunosuppressive cell populations within the tumor stroma. Targeting tumor-associated macrophages (TAM) can help to restore an anti-tumor immunity. Previously, we could show that classical TAM markers expressed in vivo need a 7 day M-CSF/dexamethasone/IL-4 (MDI) stimulation for their induction in peripheral blood monocytes (pBM) in vitro. To identify possible novel therapeutic targets on TAM, gene expression analysis of MDI-treated pBM was performed. This identified up-regulation of the purinergic G-protein coupled receptor P2Y12, the therapeutic target of the clinically approved anti-thrombotic drugs cangrelor, clopidogrel, ticagrelor, and prasugrel. We generated a peptide antibody and validated its specificity using transgenic P2Y12 + U937 cells. With the help of this antibody, P2Y12 expression was confirmed on CD68 + CD163 + TAM of melanoma in situ. Functional analysis revealed that treatment of transgenic P2Y12 + U937 cells with the receptor agonist 2-MeSADP induced ERK1/2 and Akt phosphorylation and increased the secretion of the chemokines CXCL2, CXCL7, and CXCL8. These effects could be abolished with the P2Y12 antagonist PSB0739 or with Akt and ERK inhibitors. In addition, P2Y12 + macrophages migrated towards the ADP-rich culture medium of puromycin-treated dying B16F1 melanoma cells. Cangrelor treatment blocked migration. Taken together, our results indicate that P2Y12 is an important chemotaxis receptor, which triggers migration of macrophages towards nucleotide-rich, necrotic tumor areas, and modulates the inflammatory environment upon ADP binding.

Sirtuins are NAD + -dependent deacetylases that facilitate cellular stress response. They include SirT6, which protects genome stability and regulates metabolic homeostasis through gene silencing, and whose loss induces an accelerated aging phenotype directly linked to hyperactivation of the NF-κB pathway. Here we show that SirT6 binds to the H3K9me3-specific histone methyltransferase Suv39h1 and induces monoubiquitination of conserved cysteines in the PRE-SET domain of Suv39h1. Following activation of NF-κB signaling Suv39h1 is released from the IκBα locus, subsequently repressing the NF-κB pathway. We propose that SirT6 attenuates the NF-κB pathway through IκBα upregulation via cysteine monoubiquitination and chromatin eviction of Suv39h1. We suggest a mechanism based on SirT6-mediated enhancement of a negative feedback loop that restricts the NF-κB pathway. Sirtuins are involved in the regulation of responses to diverse types of cellular stress. Here the authors describe the SirT6-dependent cysteine monoubiquitination of the histone methyltransferase Suv39h1 as part of a regulatory circuit for the NF-κB pathway.

UM is an aggressive intraocular tumor characterized by high plasticity and a propensity to metastasize in the liver. However, the underlying mechanisms governing liver tropism remain poorly understood. Given the emerging significance of exosomes, we sought to investigate the contribution of UM-derived exosomes to specific steps of the metastatic process. Firstly, we isolated exosomes from UM cells sharing a common genetic background and different metastatic properties. A comparison of protein cargo reveals an overrepresentation of proteins related to cytoskeleton remodeling and actin filament-based movement in exosomes derived from the parental cells that may favor the detachment of cells from the primary site. Secondly, we assessed the role of macrophages in reprogramming the HHSCs by exosomes. The activation of HHSCs triggered a pro-inflammatory and pro-fibrotic environment through cytokine production, upregulation of extracellular matrix molecules, and the activation of signaling pathways. Finally, we found that activated HHSCs promote increased adhesion and migration of UM cells. Our findings shed light on the pivotal role of exosomes in pre-metastatic niche construction in the liver.