Donatella Venturelli

Cellular levels of mRNA from c‐myc, c‐myb and c‐fes onc‐genes in normal myeloid anderythroid precursors of human bone marrow: an in situ hybridization study

Summary. The expression of three onc‐genes, c‐myc, c‐myb and c‐fes, has been evaluated at the cellular level in myeloid and erythroid precursors of normal human bone marrow, by ‘in situ’ hybridization with tritium‐labelled probes. A relatively large amount of m‐RNA from the three onc‐genes was detected in myeloblasts and promyelocytes, but whereas the expression of c‐myb and c‐myb decreased in more advanced stage of maturation of the myeloid lineage, c‐fes mRNA remained at a relatively high level until the granulocyte stage, c‐myc and c‐myb were expressed at a fairly high level in basophilic erythroblasts, which also showed low levels of c‐fes mRNA. No expression of these onc‐genes was detectable in more mature erythroblasts, Megakaryocytes showed high levels of m‐RNA from all three onc‐genes.

Neuroblastoma specific effects of DR-nm23 and its mutant forms on differentiation and apoptosis.

DR-nm23 belongs to a gene family which includes nm23-H1, originally identified as a candidate metastasis suppressor gene. Nm23 genes are expressed in different tumor types where their levels have been alternatively associated with reduced or increased metastatic potential. Nm23-H1, -H2, DR-nm23 and nm23-H4 all possess NDP kinase activity. Overexpression of DR-nm23 inhibits differentiation and promotes apoptosis in hematopoietic cells. By contrast, it induces morphological and biochemical changes associated with neural differentiation in neuroblastoma cells. In this study, we show that mutations in the catalytic domain and in the serine 61 phosphorylation site, possibly required for protein-protein interactions, impair the ability of DR-nm23 to induce neural differentiation. Moreover, neuroblastoma cells overexpressing wild-type or mutant DR-nm23 are less sensitive to apoptosis triggered by serum withdrawal. By subcellular fractionation, wild-type and mutant DR-nm23 localize in the cytoplasm and prevalently in the mitochondrial fraction. In co-immunoprecipitation experiments, wild-type DR-nm23 binds other members of nm23 family, but mutations in the catalytic and in the RGD domains and in serine 61 inhibit the formation of hetero-multimers. Thus, the integrity of the NDP kinase activity and the presence of a serine residue in position 61 seem essential for the ability of DR-nm23 to trigger differentiation and to bind other Nm23 proteins, but not for the anti-apoptotic effect in neuroblastoma cells. These studies underline the tissue specificity of the biological effects induced by DR-nm23 expression. Cell Death and Differentiation (2000) 7, 843–850

Expression of human c‐fes one‐gene occurs at detectable levels in myeloid but not in lymphoid cell populations

Summary. Total cellular RNA from a variety of myeloid and lymphoid cell populations, normal and leukaemic, was analysed for the expression of a human cellular one‐gene, c‐fes, by northern blot hybridization assays. The probe used was a molecularly cloned human DNA sequence homologous to the 5′ terminal sequence of v‐fes. All the myeloid cellular populations expressed the c‐fes gene. In some cell populations at an advanced stage of differentiation (circulating leucocytes from Chronic myeloid leukaemia (CML), HL60 cells induced to differentiate by retinoic acid) the level of expression was even higher than in early stages of the myeloid lineage (blast cells from AML, uninduced HL60 cells). No transcript of the c‐fes gene was detected in the different lymphoid populations studied. The occurrence of an RNA complementary to the c‐fes sequence appears sufficiently characteristic of a myeloid population to distinguish it from a lymphoid population, normal and leukaemic.

DR-nm23 expression affects neuroblastoma cell differentiation, integrin expression, and adhesion characteristics.

BACKGROUND AND PROCEDURE Nm23 gene family has been associated with metastasis suppression and differentiation. We studied DR-nm23 during neuroblastoma cells differentiation. DR-nm23 expression increased after retinoic acid induction of differentiation in human cell lines SK-N-SH and LAN-5. RESULTS In several cell lines, overexpression of DR-nm23 was associated with more differentiated phenotypes. SK-N-SH cells increased vimentin expression, increased deposition of collagen type IV, modulated integrin expression, and underwent growth arrest; the murine neuroblastoma cell line N1E-115 showed neurite outgrowth and a striking enhancement of beta1 integrin expression. Up-regulation of beta1 integrin was specifically responsible for the increase in the adhesion to collagen type I-coated plates. Finally, cells overexpressing DR-nm23 were unable to growth in soft agar. CONCLUSIONS In conclusion, DR-nm23 expression is directly involved in differentiation of neuroblastoma cells, and its ability to affects the adhesion to extracellular substrates and to inhibit growth in soft agar suggests an involvement in the metastatic potential of neuroblastoma.

Protooncogenes in the Regulation of Normal Hematopoiesis. An Antisense Approacha

Most normal circulating blood cells are highly specialized end cells that are derived from a small number of marrow progenitor cells.’.2 Progenitor cells, in turn, are derived from common pluripotent hematopoietic stem cells (PHSCs). PHSCs are capable of self-renewal, whereas committed cells have lost this potential but divide extensively during the process of differentiation.’.* The cloning of genes for several hematopoietic growth factors and the resulting production of the encoded recombinant proteins have led to major conceptual advances in understanding the regulation of normal hematopoiesis; however, intermediary metabolic events occurring within cells as steps distal to the interaction between growth factors and their own receptors are poorly understood, as are the nuclear processes that specifically regulate proliferation and differentiation of progenitor cells. In theory, activation of protooncogenes could provide a functional link between events occurring at the cellular membrane and the ensuing proliferative and maturative responses that occur in the pool of hematopoietic progenitors. The study of the regulation of hematopoietic proliferation and differentiation has primarily been hampered by difficulties in purifying a sufficient number of pure progenitor cells. In the last several years, it was shown that specific gene functions can be inhibited by exposing target cells to synthetic oligomers complementary to their ~ R N A S . ~ ” This approach eliminates the need to purify a large number of progenitor cells and provides a powerful tool for the study of the molecular basis of normal human hematopoiesis. In the following paragraphs we discuss the role of N-ras, c-abl, and c-myb (three protooncogenes encoding proteins with distinct functions) in the regulation of normal hematopoiesis.

Newborn screening for sickle cell disease in Europe: recommendations from a Pan-European Consensus Conference

Sickle Cell Disease (SCD) is an increasing global health problem and presents significant challenges to European health care systems. Newborn screening (NBS) for SCD enables early initiation of preventive measures and has contributed to a reduction in childhood mortality from SCD. Policies and methodologies for NBS vary in different countries, and this might have consequences for the quality of care and clinical outcomes for SCD across Europe. A two‐day Pan‐European consensus conference was held in Berlin in April 2017 in order to appraise the current status of NBS for SCD and to develop consensus‐based statements on indications and methodology for NBS for SCD in Europe. More than 50 SCD experts from 13 European countries participated in the conference. This paper aims to summarise the discussions and present consensus recommendations which can be used to support the development of NBS programmes in European countries where they do not yet exist, and to review existing programmes.