PLC-beta 1 regulates the expression of miR-210 during mithramycin-mediated erythroid differentiation in K562 cells

PLC-beta 1 (PLCβ1) inhibits erythroid differentiation induced by mithramycin (MTH) by targeting miR-210 expression. MicroRNA-210 (miR-210) has been reported to be upregulated in various types of human malignancy suggesting that it has an important role in tumorigenesis. Inhibition of miR-210 affects the erythroid differentiation pathway and it occurs to a greater extent in MTH-treated cells. In this paper we have analyzed the effect of MTH on human K562 cells differentiation. Overexpression of PLCβ1 suppresses the differentiation of K562 elicited by MTH as demonstrated by the absence of γ-globin expression. Inhibition of PLCβ1 expression is capable to promote the differentiation process leading to a recovery of γ-globin gene even in the absence of MTH. Our experimental evidences suggest that PLCβ1 signalling regulates erythropoiesis through miR-210. Indeed overexpression of PLCβ1 leads to a decrease of miR-210 expression after MTH treatment. Moreover miR-210 is up-regulated through both proliferation and differentiation events when PLCβ1 expression is down-regulated. Therefore we suggest a novel role for PLCβ1 in regulating miR-210 and our data hint at the fact that, in human K562 erythroleukemia cells, the modulation of PLCβ1 expression is able to exert an impairment of normal erythropoiesis as assessed by γ-globin expression

K562 cell proliferation is modulated by PLCβ1 through a PKCα-mediated pathway

Phospholipase C β1 (PLCβ1) is known to play an important role in cell proliferation. Previous studies reported aninvolvement of PLCβ1 in G0-G1/S transition and G2/M progression in Friend murine erythroleukemia cells (FELC). However,little has been found about its role in human models. Here, we used K562 cell line as human homologous of FELC inorder to investigate the possible key regulatory role of PLCβ1 during cell proliferation of this humancell line. Our studies on the effects of the overexpression of both these isoforms showed a specific and positive connection between cyclinD3 and PLCβ1 in K562 cells, which led to a prolonged S phase of the cell cycle and a delay in cell proliferation. In order to shed light on this mechanism, we decided to study the possible involvement of protein kinases C (PKC), known to be direct targets of PLC signaling and important regulators of cell proliferation. Our data showed a peculiar decrease of PKCα levels in cells overexpressing PLCβ1. Moreover, when we silenced PKCα, by RNAi technique, in order to mimic the effects of PLCβ1, we caused the same upregulation of cyclin D3 levels and the same decrease of cell proliferation found in PLCβ1-overexpressing cells. The key features emerging from our studies in K562 cells is that PLCβ1 targets cyclin D3, likely through a PKCα-mediated-pathway, and that, as a downstream effect of its activity, K562 cells undergo an accumulation in the S phase of the cell cycle

PLC-beta 1 regulates the expression of miR-210 during mithramycin-mediated erythroid differentiation in K562 cells

PLC-beta 1 (PLCβ1) inhibits in human K562 cells erythroid differentiation induced by mithramycin (MTH) by targeting miR-210 expression. Inhibition of miR-210 affects the erythroid differentiation pathway and it occurs to a greater extent in MTH-treated cells. Overexpression of PLCβ1 suppresses the differentiation of K562 elicited by MTH as demonstrated by the absence of γ-globin expression. Inhibition of PLCβ1 expression is capable to promote the differentiation process leading to a recovery of γ-globin gene even in the absence of MTH. Our experimental evidences suggest that PLCβ1 signaling regulates erythropoiesis through miR-210. Indeed overexpression of PLCβ1 leads to a decrease of miR-210 expression after MTH treatment. Moreover miR-210 is up-regulated when PLCβ1 expression is down-regulated. When we silenced PKCα by RNAi technique, we found a decrease in miR-210 and γ-globin expression levels, which led to a severe slowdown of cell differentiation in K562 cells and these effects were the same encountered in cells overexpressing PLCβ1. Therefore we suggest a novel role for PLCβ1 in regulating miR-210 and our data hint at the fact that, in human K562 erythroleukemia cells, the modulation of PLCβ1 expression is able to exert an impairment of normal erythropoiesis as assessed by γ-globin expression

BMP-2 induced expression of PLC beta1 that is a positive regulator of osteoblast differentiation

C2C12 is an immortalized mouse myoblast cell line. The cells readily proliferate in high-serum conditions, and differentiate and fuse in low-serum conditions. While this cell line is a very useful tool to study aspects of myogenesis, metabolism and muscle biology, however, treatment of C2C12 cells with bone morphogenic protein (BMPs) induces cells to differentiate into osteoblasts. Osteoblast differentiation is controlled by diversified signaling proteins and transcription factors, essentially BMP-2, Osterix (Osx/Sp7) and Runx2, finally associating with the expression of late osteoblast marker genes, like ALPL and Bglap. These peculiarities make C2C12 progenitor cells a skillful prototype to investigate the molecular mechanism that control cell destiny specification and terminal differentiation. In the current investigation, we took improvement of the differentiation peculiarities of the mouse C2C12 cell line to analyze whether changes in PLCbeta1 expression and its nuclear localization might regulate or affect their terminal osteogenic differentiation. We demonstrated that overexpression of PLCβ1 enhances the osteogenic differentiation of C2C12 elicited by BMP-2 as demonstrated by the presence of osteoblast marker genes expression. In the present study we also showed that miR-214 suppressed osteogenic differentiation through the regulation of nuclear PLCβ1 by targeting Osterix

A Role for Nuclear Phospholipase Cβ1 in Cell Cycle Control

Phosphoinositide signaling resides in the nucleus, and among the enzymes of the cycle, phospholipase C (PLC) appears as the key element both in Saccharomyces cerevisiae and in mammalian cells. The yeast PLC pathway produces multiple inositol polyphosphates that modulate distinct nuclear processes. The mammalian PLCbeta(1), which localizes in the nucleus, is activated in insulin-like growth factor 1-mediated mitogenesis and undergoes down-regulation during murine erythroleukemia differentiation. PLCbeta(1) exists as two polypeptides of 150 and 140 kDa generated from a single gene by alternative RNA splicing, both of them containing in the COOH-terminal tail a cluster of lysine residues responsible for nuclear localization. These clues prompted us to try to establish the critical nuclear target(s) of PLCbeta(1) subtypes in the control of cell cycle progression. The results reveal that the two subtypes of PLCbeta(1) that localize in the nucleus induce cell cycle progression in Friend erythroleukemia cells. In fact when they are overexpressed in the nucleus, cyclin D3, along with its kinase (cdk4) but not cyclin E is overexpressed even though cells are serum-starved. As a consequence of this enforced expression, retinoblastoma protein is phosphorylated and E2F-1 transcription factor is activated as well. On the whole the results reveal a direct effect of nuclear PLCbeta(1) signaling in G(1) progression by means of a specific target, i.e. cyclin D3/cdk4

Nuclear phospholipase Cβ1 interactome: a morphological and proteomic approach

Inositide-dependent phospholipase Cβ1 (PI-PLCβ1b) has two isoforms generated by alternative splicing (PI-PLCβ1a and PI-PLCβ1b). In murine erythroleukemia (MEL) cells both the isoforms are present within the nucleus, but PI-PLCβ1b is exclusively nuclear. Our group has demonstrated that PI-PLCβ1 nuclear localisation is crucial for its function, although the mechanism by which PI-PLCβ1 is imported into the nucleus has never been carefully investigated. The purpose of the present study was to get more insights on the protein interactome of PI-PLCβ1b, namely the proteins present in the nucleus. Immuno-affinity purification coupled with tandem mass spectrometry analysis have been used to purify and identify PI-PLCβ1b interaction binding partners from Friend’s erythroleukemia isolated nuclei. Gene ontology and protein-protein interaction network were performed to analyze data. Some interactions were already characterized, such as the binding with the splicing factor SRp20 and the lamin B. Among the proteins identified, the binding of eEF1A and prohibitin 2 with PI-PLCβ1b was confirmed by western blot analysis. Of particular interest was the identification of importin a, importin b1 and Ran, which interact with PI-PLCβ1b. These proteins are believed to be involved in the import mechanism from the cytoplasm to the nucleus. Further analysis by overexpressing both wild type and cytoplasmatic mutant of PI-PLCβ1, suggests that importin b1 is responsible for the localisation of PI-PLCβ1b in the nucleus, giving new insight into the mechanism of trafficking of this signaling molecule

The Italian Earthquakes and Tsunami Monitoring and Surveillance Systems

The Osservatorio Nazionale Terremoti (ONT) is the Italian seismic operational centre for monitoring earthquake, it is part of Istituto Nazionale di Geofisica e Vulcanologia (INGV) the largest Italian research institution, with focus in Earth Sciences. INGV runs the Italian National Seismic Network (network code IV) and other networks at national scale for monitoring earthquakes and tsunami. INGV is a primary node of European Integrated Data Archive (EIDA) for archiving and distributing, continuous, quality checked seismic waveforms (strong motion and weak motion recordings). ONT designed the data acquisition system to accomplish, in near-real-time, automatic earthquake detection, hypocentre and magnitude determination and evaluation of moment tensors, shake maps and other products. Database archiving of all parametric results are closely linked to the existing procedures of the INGV seismic monitoring environment and surveillance procedures. ONT organize the Italian earthquake surveillance service and the tsunami alert service (INGV is Tsunami Service Provider of the ICG/NEAM for the entire Mediterranean basin). We provide information to the Dipartimento di Protezione Civile (DPC) and to several Mediterranean countries. Earthquakes information are revised routinely by the analysts of the Italian Seismic Bulletin. The results are published on the web and are available to the scientific community and the general public.PublishedMontreal1SR TERREMOTI - Sorveglianza Sismica e Allerta Tsunam

Low in‑hospital mortality rate in patients with COVID‑19 receiving thromboprophylaxis: data from the multicentre observational START‑COVID Register

Abstract COVID-19 infection causes respiratory pathology with severe interstitial pneumonia and extra-pulmonary complications; in particular, it may predispose to thromboembolic disease. The current guidelines recommend the use of thromboprophylaxis in patients with COVID-19, however, the optimal heparin dosage treatment is not well-established. We conducted a multicentre, Italian, retrospective, observational study on COVID-19 patients admitted to ordinary wards, to describe clinical characteristic of patients at admission, bleeding and thrombotic events occurring during hospital stay. The strategies used for thromboprophylaxis and its role on patient outcome were, also, described. 1091 patients hospitalized were included in the START-COVID-19 Register. During hospital stay, 769 (70.7%) patients were treated with antithrombotic drugs: low molecular weight heparin (the great majority enoxaparin), fondaparinux, or unfractioned heparin. These patients were more frequently affected by comorbidities, such as hypertension, atrial fibrillation, previous thromboembolism, neurological disease,and cancer with respect to patients who did not receive thromboprophylaxis. During hospital stay, 1.2% patients had a major bleeding event. All patients were treated with antithrombotic drugs; 5.4%, had venous thromboembolism [30.5% deep vein thrombosis (DVT), 66.1% pulmonary embolism (PE), and 3.4% patients had DVT + PE]. In our cohort the mortality rate was 18.3%. Heparin use was independently associated with survival in patients aged ≥ 59 years at multivariable analysis. We confirmed the high mortality rate of COVID-19 in hospitalized patients in ordinary wards. Treatment with antithrombotic drugs is significantly associated with a reduction of mortality rates especially in patients older than 59 years