Multivalency of nucleosome recognition by LEDGF.

Eukaryotic transcription is dependent on specific histone modifications. Their recognition by chromatin readers triggers complex processes relying on the coordinated association of transcription regulatory factors. Although various modification states of a particular histone residue often lead to differential outcomes, it is not entirely clear how they are discriminated. Moreover, the contribution of intrinsically disordered regions outside of the specialized reader domains to nucleosome binding remains unexplored. Here, we report the structures of a PWWP domain from transcriptional coactivator LEDGF in complex with the H3K36 di- and trimethylated nucleosome, indicating that both methylation marks are recognized by PWWP in a highly conserved manner. We identify a unique secondary interaction site for the PWWP domain at the interface between the acidic patch and nucleosomal DNA that might contribute to an H3K36-methylation independent role of LEDGF. We reveal DNA interacting motifs in the intrinsically disordered region of LEDGF that discriminate between the intra- or extranucleosomal DNA but remain dynamic in the context of dinucleosomes. The interplay between the LEDGF H3K36-methylation reader and protein binding module mediated by multivalent interactions of the intrinsically disordered linker with chromatin might help direct the elongation machinery to the vicinity of RNA polymerase II, thereby facilitating productive elongation.

Nestin expression throughout multistep pathogenesis of multiple myeloma.

The stem cell marker nestin (NES) is found in dividing cells of developing and regenerating tissues. Upon terminal differentiation, NES expression is diminished but may be re-expressed following injury or in cancer. Surprisingly, we recently confirmed NES as a tumour-specific marker for mature CD138(+) 38(+) plasma cells (PC) in multiple myeloma (MM). The present study analysed NES expression throughout the spectrum of MM developmental stages, starting with individuals with no haematological malignancy, through monoclonal gammopathy of undetermined significance (MGUS) and MM to plasma cell leukaemia (PCL) and MM cell lines. NES was analysed in bone marrow PC of 163 MM, four PCL and nine MGUS patients, 10 individuals with no haematological malignancy and 6 myeloma cell lines (OPM-2, RPMI-8226, MOLP-8, U-266, EJM, NCI-H929) by flow cytometry and/or real-time polymerase chain reaction or immunochemistry. We observed a tendency of increased NES expression in parallel with disease progression. NES was evaluated as a reliable marker for accurate discrimination between MM patients and the control group. High NES levels were strongly associated with the presence of 1q21 gain. For the first time, NES was demonstrated to predict worse response to conventional therapy/novel agents. These results suggest that NES might become a useful clinical parameter with an important role in MM pathogenesis.

Myeloma stem cell concepts, heterogeneity and plasticity of multiple myeloma.

Multiple myeloma (MM) is a haematological malignancy characterized by the accumulation of clonal plasma cells (PCs) in the bone marrow (BM). Although novel therapeutic strategies have prolonged survival of patients, the disease remains difficult to treat with a high risk of relapse. The failure of therapy is thought to be associated with a persistent population of the so-called MM stem cells or myeloma initiating cells (MIC) that exhibit tumour-initiating potential, self-renewal and resistance to chemotherapy. However, the population responsible for the origin and sustainability of tumour mass has not been clearly characterized so far. This review summarizes current myeloma stem cell concepts and suggests that high phenotypic and intra-clonal heterogeneity, together with plasticity potential of MM might be other contributing factors explaining discrepancies among particular concepts and contributing to the treatment failure.

Association study of selected genetic polymorphisms and occurrence of venous thromboembolism in patients with multiple myeloma who were treated with thalidomide.

INTRODUCTION/BACKGROUND: Venous thromboembolism (VTE), with the subsequent risk of pulmonary embolism, is a common adverse effect of thalidomide treatment in patients with multiple myeloma (MM). In our retrospective study, we analyzed candidate single-nucleotide polymorphisms (SNP), CINP (rs7011), CETP (rs289747), ALDH1A1 (rs610529), CDKN1A (rs3829963), GAN (rs2608555), vascular endothelial growth factor (rs699947), and ALDH1A1 (rs168351), previously identified in a large association study based on the hypothesis-driven candidate gene approach nominated by the International Myeloma Foundation "Bank On A Cure" (3404 SNPs). In that study, the researchers built a classification tree that enables prediction of individual risk of VTE in patients with MM. PATIENTS AND METHODS: Genotypes of these SNPs were determined in an independent cohort of 111 patients with MM through TaqMan real-time polymerase chain reaction (PCR) allelic discrimination and were used for prediction of individual VTE risk. RESULTS: The results of this study did not confirm the ability of this classification tree to predict VTE risk in patients with MM from the Czech Republic; of these patients, 21 (19%) developed high-grade VTE. However, in patients with VTE, we found higher frequency of the AC genotype in the CDKN1A gene (42.9% vs. 16.7%; odds ratio 3.64) in comparison with the CC genotype (P = .015). SNPs of other genes as well as age and sex of the patients had no statistically significant influence on the risk of VTE. CONCLUSION: Further studies are needed to confirm the initial analysis that provided predictive information of genetic variations in patients with myeloma that may influence risk of VTE.

Levels of angiogenic factors in patients with multiple myeloma correlate with treatment response.

Angiogenesis plays a significant role in the pathogenesis of multiple myeloma (MM). We have measured concentrations of angiogenesis activators, including vascular endothelial growth factor (VEGF), basic fibroblast growth factor, and hepatocyte growth factor (HGF), and inhibitors, including endostatin, thrombospondin-1 (TSP-1), and angiostatin in the peripheral and bone marrow blood of MM patients at diagnosis and after high-dose chemotherapy. We have analyzed 96 patients with secretory MM. Serial measurements of angiogenesis factors/inhibitors were analyzed in the plasma by subgroups based on the best treatment response. Concentrations of angiogenic factors were determined in the peripheral blood and bone marrow plasma. There were significant decreases of VEGF and HGF levels and a significant increase in TSP-1 concentrations in the bone marrow plasma of patients who achieved complete or very good partial response in contrast to those who had partial or no response. VEGF and HGF levels decrease but those of TSP-1 increase after successful treatment for MM, indicating a reduction in the rate of angiogenesis.

Nestin expression in the cell lines derived from glioblastoma multiforme.

BACKGROUND: Nestin is a protein belonging to class VI of intermediate filaments that is produced in stem/progenitor cells in the mammalian CNS during development and is consecutively replaced by other intermediate filament proteins (neurofilaments, GFAP). Down-regulated nestin may be re-expressed in the adult organism under certain pathological conditions (brain injury, ischemia, inflammation, neoplastic transformation). Our work focused on a detailed study of the nestin cytoskeleton in cell lines derived from glioblastoma multiforme, because re-expression of nestin together with down-regulation of GFAP has been previously reported in this type of brain tumor. METHODS: Two cell lines were derived from the tumor tissue of patients treated for glioblastoma multiforme. Nestin and other cytoskeletal proteins were visualized using imunocytochemical methods: indirect immunofluorescence and immunogold-labelling. RESULTS: Using epifluorescence and confocal microscopy, we described the morphology of nestin-positive intermediate filaments in glioblastoma cells of both primary cultures and the derived cell lines, as well as the reorganization of nestin during mitosis. Our most important result came through transmission electron microscopy and provided clear evidence that nestin is present in the cell nucleus. CONCLUSION: Detailed information concerning the pattern of the nestin cytoskeleton in glioblastoma cell lines and especially the demonstration of nestin in the nucleus represent an important background for further studies of nestin re-expression in relationship to tumor malignancy and invasive potential.