Physiological expression of miR-130a during differentiation of CD34+ human hematopoietic stem cells results in the inhibition of monocyte differentiation.

MicroRNAs (miRNA) are small noncoding RNAs that regulate gene expression by targeting mRNAs in a sequence specific manner, thereby determining their degradation or inhibiting translation. They are involved in processes such as proliferation, differentiation and apoptosis by fine-tuning the expression of genes underlying such events. The expression of specific miRNAs is involved in hematopoietic differentiation and their deregulation contributes to the development of hematopoietic malignancies such as acute myeloid leukemia (AML). miR-130a is over-expressed in AML. Here we show that miR-130a is physiologically expressed in myeloblasts and down-regulated during monocyte differentiation. Gain- and loss-of-function experiments performed on CD34+ human hematopoietic stem cells confirmed that expression of miR-130a inhibits monocyte differentiation by interfering with the expression of key transcription factors HOXA10, IRF8, KLF4, MAFB and PU-1. The data obtained in this study highlight that the correct modulation of miR-130a is necessary for normal differentiation to occur and confirming that deregulation of this miRNA might underlie the differentiation block occurring in AML.

Magnesium Is a Key Regulator of the Balance between Osteoclast and Osteoblast Differentiation in the Presence of Vitamin D3.

Magnesium (Mg) is crucial for bone health. Low concentrations of Mg inhibit the activity of osteoblasts while promoting that of osteoclasts, with the final result of inducing osteopenia. Conversely, little is known about the effects of high concentrations of extracellular Mg on osteoclasts and osteoblasts. Since the differentiation and activation of these cells is coordinated by vitamin D3 (VD3), we investigated the effects of high extracellular Mg, as well as its impact on VD3 activity, in these cells. U937 cells were induced to osteoclastic differentiation by VD3 in the presence of supra-physiological concentrations (>1 mM) of extracellular Mg. The effect of high Mg concentrations was also studied in human bone-marrow-derived mesenchymal stem cells (bMSCs) induced to differentiate into osteoblasts by VD3. We demonstrate that high extra-cellular Mg levels potentiate VD3-induced osteoclastic differentiation, while decreasing osteoblastogenesis. We hypothesize that Mg might reprogram VD3 activity on bone remodeling, causing an unbalanced activation of osteoclasts and osteoblasts.

Chromosome positioning in interphase nuclei of hematopoietic stem cell and myeloid precursor.

Human myelopoiesis is an intriguing biological process during which multipotent stem cells limit their differentiation potential generating precursors that evolve into terminally differentiated cells. The differentiation process is correlated with differential gene expression and changes in nuclear architecture. In interphase, chromosomes are distinct entities known as chromosome territories and they show a radial localization that could result in a constrain of inter-homologous distance. This element plays a role in genome stability and gene expression. Here, we provide the first experimental evidence of 3D chromosomal arrangement considering two steps of human normal myelopoiesis. Specifically, multicolor 3D-FISH and 3D image analysis revealed that, in both normal human hematopoietic stem cells and myelod precursors CD14-, chromosomal position is correlated with gene density. However, we observed that inter-homologue distances are totally different during differentiation. This could be associated with differential gene expression that we found comparing the two cell types. Our results disclose an unprecedented framework relevant for deciphering the genomic mechanisms at the base of normal human myelopoiesis.

KLF4 Mediates the Effect of 5-ASA on the ß-Catenin Pathway in Colon Cancer Cells.

Mesalazine (5-ASA) is an aminosalicylate anti-inflammatory drug capable of inducing µ-protocadherin, a protein expressed by colorectal epithelial cells that is downregulated upon malignant transformation. Treatment with 5-ASA restores µ-protocadherin expression and promotes the sequestration of ß-catenin to the plasma membrane. Here, we show that 5-ASA-induced µ-protocadherin expression is directly regulated by the KLF4 transcription factor. In addition, we suggest the existence of a dual mechanism whereby 5-ASA-mediated ß-catenin inhibition is caused by µ-protocadherin-dependent sequestration of ß-catenin to the plasma membrane and by the direct binding of KLF4 to ß-catenin. In addition, we found that 5-ASA treatment suppresses the expression of miR-130a and miR-135b, which target KLF4 mRNA, raising the possibility that this mechanism is involved in the increased expression of KLF4 induced by 5-ASA. Cancer Prev Res; 11(8); 503-10. ©2018 AACR.

Epigenetically induced ectopic expression of UNCX impairs the proliferation and differentiation of myeloid cells.

We here describe a leukemogenic role of the homeobox gene UNCX, activated by epigenetic modifications in acute myeloid leukemia (AML). We found the ectopic activation of UNCX in a leukemia patient harboring a t(7;10)(p22;p14) translocation, in 22 of 61 of additional cases [a total of 23 positive patients out of 62 (37.1%)], and in 6 of 75 (8%) of AML cell lines. UNCX is embedded within a low-methylation region (canyon) and encodes for a transcription factor involved in somitogenesis and neurogenesis, with specific expression in the eye, brain, and kidney. UNCX expression turned out to be associated, and significantly correlated, with DNA methylation increase at its canyon borders based on data in our patients and in archived data of patients from The Cancer Genome Atlas. UNCX-positive and -negative patients displayed significant differences in their gene expression profiles. An enrichment of genes involved in cell proliferation and differentiation, such as MAP2K1 and CCNA1, was revealed. Similar results were obtained in UNCX-transduced CD34+ cells, associated with low proliferation and differentiation arrest. Accordingly, we showed that UNCX expression characterizes leukemia cells at their early stage of differentiation, mainly M2 and M3 subtypes carrying wild-type NPM1 We also observed that UNCX expression significantly associates with an increased frequency of acute promyelocytic leukemia with PML-RARA and AML with t(8;21)(q22;q22.1); RUNX1-RUNX1T1 classes, according to the World Health Organization disease classification. In summary, our findings suggest a novel leukemogenic role of UNCX, associated with epigenetic modifications and with impaired cell proliferation and differentiation in AML.

Remodeling of nuclear landscapes during human myelopoietic cell differentiation maintains co-aligned active and inactive nuclear compartments.

BACKGROUND: Previous studies of higher order chromatin organization in nuclei of mammalian species revealed both structural consistency and species-specific differences between cell lines and during early embryonic development. Here, we extended our studies to nuclear landscapes in the human myelopoietic lineage representing a somatic cell differentiation system. Our longterm goal is a search for structural features of nuclei, which are restricted to certain cell types/species, as compared to features, which are evolutionary highly conserved, arguing for their basic functional roles in nuclear organization. RESULTS: Common human hematopoietic progenitors, myeloid precursor cells, differentiated monocytes and granulocytes analyzed by super-resolution fluorescence microscopy and electron microscopy revealed profound differences with respect to global chromatin arrangements, the nuclear space occupied by the interchromatin compartment and the distribution of nuclear pores. In contrast, we noted a consistent organization in all cell types with regard to two co-aligned networks, an active (ANC) and an inactive (INC) nuclear compartment delineated by functionally relevant hallmarks. The ANC is enriched in active RNA polymerase II, splicing speckles and histone signatures for transcriptionally competent chromatin (H3K4me3), whereas the INC carries marks for repressed chromatin (H3K9me3). CONCLUSIONS: Our findings substantiate the conservation of the recently published ANC-INC network model of mammalian nuclear organization during human myelopoiesis irrespective of profound changes of the global nuclear architecture observed during this differentiation process. According to this model, two spatially co-aligned and functionally interacting active and inactive nuclear compartments (ANC and INC) pervade the nuclear space.