Unknown,Transcriptomics,Genomics,Proteomics

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Bone morphogenetic protein-7 is a MYC target with pro-survival functions in childhood medulloblastoma

ABSTRACT: Medulloblastoma (MB) is the most common malignant brain tumor in children, among whom overexpression or amplification of MYC oncogenes has been associated with poor clinical outcome. Although the MYC functions during normal development and oncogenesis in various systems have been extensively investigated, the transcriptional targets mediating MYC effects in MB are still elusive. Their identification and roles during MB onset and progression are important and will ultimately suggest novel potential therapeutic targets. cDNA microarray analysis was used to compare the effects of overexpressing and silencing MYC on the transcriptome of a MB-derived cell line. We identified 209 genes with potential relevance to MYC-dependent cellular responses in MB. Among the MYC-responsive genes, we found members of the bone morphogenetic protein (BMP) signaling pathway, which plays a crucial role during the development of the cerebellum. In particular, the cytokine gene BMP7 was identified as a direct target of MYC in MB cells. Similar to the effect induced by BMP7 silencing by siRNA, the use of a small-molecule inhibitor of the BMP/SMAD signaling pathway reduced cell viability in a panel of MB cells. Altogether, our findings indicate that high MYC levels drive BMP7 expression in MB to induce pro-survival and pro-proliferative cellular pathways. This observation suggests that targeting the BMP/SMAD pathway may be a new therapeutic concept for the treatment of childhood MB. 6 samples (3 replicates for each sample): wild-type MB cell line, empty vector-transfected control cell line, 2 c-MYC-overexpressing clones, 1 clone upon c-MYC silencing, 1 silencing control cell line.

ORGANISM(S): Homo sapiens

SUBMITTER: Stefan Zoller

PROVIDER: E-GEOD-22139 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Bone morphogenetic protein-7 is a MYC target with prosurvival functions in childhood medulloblastoma.

Fiaschetti G G Castelletti D D Zoller S S Schramm A A Schroeder C C Nagaishi M M Stearns D D Mittelbronn M M Eggert A A Westermann F F Ohgaki H H Shalaby T T Pruschy M M Arcaro A A Grotzer M A MA

Oncogene 20110214 25

Medulloblastoma (MB) is the most common malignant brain tumor in children. It is known that overexpression and/or amplification of the MYC oncogene is associated with poor clinical outcome, but the molecular mechanisms and the MYC downstream effectors in MB remain still elusive. Besides contributing to elucidate how progression of MB takes place, most importantly, the identification of novel MYC-target genes will suggest novel candidates for targeted therapy in MB. A group of 209 MYC-responsive ...[more]

PMID: 21317922

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Project description:Background: MYC is a transcription factor encoded by the c-MYC gene (thereafter termed MYC). MYC is key transcription factor involved in many central cellular processes including ribosomal biogenesis. MYC is overexpressed in the majority of human tumours including aggressive B-cell lymphoma especially Burkitt's lymphoma. Although Burkitt's lymphoma is a highlight example for MYC overexpression due to a chromosomal translocation, no global analysis of MYC binding sites by chromatin immunoprecipitation (ChIP) followed by global next generation sequencing (ChIP-Seq) has been conducted so far in Burkitt's lymphoma. Methodology/Principal Findings: ChIP-Seq was performed with a MYC-specific antibody giving rise to 7,054 predicted MYC binding sites after bioinformatics analysis of a total of 19 million sequence reads. In line with previous findings, binding sites accumulate in gene sets known to be involved in the ribosomal biogenesis, histone acetyltransferase and methyltransferase complexes and the cell cycle demonstrating a regulatory role of MYC in these processes. Unexpectedly, MYC binding sites also accumulate in genes typically expressed in mature B-cells. To assess the functional consequences of altered MYC binding, the ChIP-Seq data were supplemented with siRNA mediated knock-downs of MYC in BL cell lines followed by gene expression profiling. Interestingly, amongst others, genes involved in B-cell function were up-regulated in response to MYC silencing. Conclusion/Significance: The 7,054 MYC-binding sites identified by our ChIP-Seq approach greatly extend the knowledge regarding MYC binding in Burkitt's lymphoma and sheds further light on the enormous complexity of the MYC regulatory network. Especially our observation that (i) many B-cell relevant genes are targeted by MYC and (ii) that MYC down-regulation leads to an up-regulation of B-cell genes highlights an interesting aspect of BurkittM-BM-4s lymphoma biology. [ChIP-Seq] Analysis of MYC DNA binding sites by ChiP-Seq in 5 BurkittM-BM-4s lymphoma cell lines (Raji, Ramos, Blue1, BL41, CA46) [mRNA expression profiling] siRNA-mediated knock-down of MYC was done employing the BL cell lines Raji, BL41 and Blue1 in order to detect MYC-driven gene expression changes. For this purpose, the cells were Amaxa-transfected using MYC smart pool siRNA and control siRNA (Thermo Scientific/Dharmacon, Erembodegem, Belgium), respectively.

Project description:Medulloblastomas (MBs) are cerebellar tumors that can be classified into molecularly distinct subgroups that differ in pathology and prognosis. The mechanisms that underlie subgroup specification are largely unknown. While human SHH MBs express MYCN, Group3 (G3) MBs are associated with c-MYC (MYC) overexpression and often show metastasis that confers a poor prognosis. Although MYC proteins are thought to be functionally exchangeable, ectopic expression of Myc or N-myc in Trp53-/-;Cdkn2c-/- cerebellar granule neuron progenitors (GNPs) induces G3 and SHH MBs, respectively, demonstrating that each Myc protein has distinct biological properties. We now show that Myc and N-myc differ in their affinity to Miz1 and that Myc, but not N-myc, effectively recruits Miz1 to its target sites on chromatin. The interaction of Myc with Miz1 is required for the genesis of G3 MB. Myc suppresses ciliogenesis and âreprogramsâ the transcriptome of SHH-dependent GNPs to stem-like cells by repressing genes highly expressed in SHH MB via Miz1. Consistently, target genes of Myc/Miz1 are repressed in human G3 MBs but not in other MB subgroups. Collectively, the data show that the interaction of Myc with Miz1 is a defining hallmark of G3 MB development. In this study, we show that Myc and N-myc differ in their affinity for Miz1, and Myc/Miz1 interaction is required for Group3 medulloblastoma (MB). The gene expression profiles of these tumors were compared to our previously published Group3 MB model as well as SHH model of MB (Kawachi et al., 2012, Cancer Cell). Cerebellar granule neuron progenitors (GNPs) [dka220-222] from postnatal (P) 6 Math1-GFP;Trp53-/-;Cdkn2c-/-. For SHH medulloblastomas, [dka204-206] and [blm015-017 or dka050-dka051], spontaneous medulloblastomas from Cdkn2c-/-;Trp53Fl/Fl;Nestin-Cre (Kawachi et al., 2012, Cancer Cell) and Ptch1+/-;Cdkn2c-/- (Uziel et al., 2005, Genes Dev) were used, respectively. Myc- [dka201-203] and MycV394D (termed MycVD thereafter)- [bvo017-bvo023] tumors were from Trp53-/-;Cdkn2c-/- cerebella of P6-P7 pups. Myc/ÎPOZ tumors [bvo002-bvo006] were obtained from Trp5Fl/Fl;Miz1ÎPOZ/POZ;Nestin-Cre cerebella of P6-P7 pups.

Project description:Medulloblastoma (MB) is the most common malignant brain tumor. MB is a cerebellar tumor that occurs mostly in children between the ages of 3-7 years but also in adults. Human MBs are classified into four subgroups: Wingless (WNT), Sonic Hedgehog (SHH), Group 3 (G3) and G4, each of which with distinct molecular signatures. SHH MBs with MYCN amplification and TP53 mutations and G3 MBs characterized by C-MYC (MYC) overexpression in ~17% of cases from gene amplification with stem like properties, are the most aggressive and least curable with current therapeutic regimen. Using an orthotopic transplant approach, we found that enforced expression of MYCN in cerebellar granule neural progenitors (CGNPs) from 5-7 days old Trp53-null mice induced SHH MBs after transfer in the cortices or cerebella of naive recipient mice. In contrast, overexpression of MYC induced G3 MBs. Because MYCN and MYC bind to the same E-box DNA sequences, we hypothesized that the difference between MYC and MYCN-induced gene expression and tumor identity might be due to their interaction with different partners in CGNPs. In this study, we investigated the role of the Myc interacting zinc finger protein 1 (MIZ1). We found that MIZ1 binds with higher affinity to MYC than to MYCN and that MIZ1 and MYC co-occupy thousands of promoters in G3 MB. Remarkably, enforced expression of a MYC mutant (MYCV394D) that no longer binds to MIZ1 in Trp53-null CGNPs or expression of wild type MYC in CGNPs that lack functional Miz1 resulted in massive changes of the resulting tumorâ??s transcriptional program. Tumors differed from both SHH and G3 medulloblastoma and had a later time of onset compared to MYC-driven G3 medulloblastoma. Our data demonstrate that interaction of MYC with MIZ1 is required for the development of G3 medulloblastoma. ChIP-Seq experiments for Miz1, c-Myc and NMyc from murine medulloblastoma material. Either sorted tumor cells (Miz1 and c-Myc) or spheres from tumor cells (Miz1 and NMyc) were used. Input-samples were sequenced as controls. RNA-Seq from G3 medulloblastomas after restoration of Atoh1 expression.

Dataset Information

Bone morphogenetic protein-7 is a MYC target with pro-survival functions in childhood medulloblastoma

Publications

Bone morphogenetic protein-7 is a MYC target with prosurvival functions in childhood medulloblastoma.

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