Cotterman R, Knoepfler PS.. N-Myc regulates expression of pluripotency genes in neuroblastoma including lif, klf2, klf4, and lin28b. PLoS One 4: e5799

Department of Cell Biology and Human Anatomy, and Stem Cell Program, University of California Davis School of Medicine, Shriners Hospital For Children Northern California, Sacramento, California, USA.
PLoS ONE (Impact Factor: 3.23). 02/2009; 4(6):e5799. DOI: 10.1371/journal.pone.0005799
Source: PubMed


myc genes are best known for causing tumors when overexpressed, but recent studies suggest endogenous myc regulates pluripotency and self-renewal of stem cells. For example, N-myc is associated with a number of tumors including neuroblastoma, but also plays a central role in the function of normal neural stem and precursor cells (NSC). Both c- and N-myc also enhance the production of induced pluripotent stem cells (iPSC) and are linked to neural tumor stem cells. The mechanisms by which myc regulates normal and neoplastic stem-related functions remain largely open questions. Here from a global, unbiased search for N-Myc bound genes using ChIP-chip assays in neuroblastoma, we found lif as a putative N-Myc bound gene with a number of strong N-Myc binding peaks in the promoter region enriched for E-boxes. Amongst putative N-Myc target genes in expression microarray studies in neuroblastoma we also found lif and three additional important embryonic stem cell (ESC)-related factors that are linked to production of iPSC: klf2, klf4, and lin28b. To examine the regulation of these genes by N-Myc, we measured their expression using neuroblastoma cells that contain a Tet-regulatable N-myc transgene (TET21N) as well as NSC with a nestin-cre driven N-myc knockout. N-myc levels closely correlated with the expression of all of these genes in neuroblastoma and all but lif in NSC. Direct ChIP assays also indicate that N-Myc directly binds the lif promoter. N-Myc regulates trimethylation of lysine 4 of histone H3 in the promoter of lif and possibly in the promoters of several other stem-related genes. Together these findings indicate that N-Myc regulates overlapping stem-related gene expression programs in neuroblastoma and NSC, supporting a novel model by which amplification of the N-myc gene may drive formation of neuroblastoma. They also suggest mechanisms by which Myc proteins more generally contribute to maintenance of pluripotency and self-renewal of ESC as well as to iPSC formation.

14 Reads
  • Source
    • "Despite numerous genes demonstrating co-regulation with MYCN, few have been definitively demonstrated to be direct MYCN targets. These include ODC1, PTMA, IGF1R and hTERT [7] [12] [13] and more recently MDM2, CRABP-II, DKK1, LIF, KLF2, KLF4 and LIN28b [14] [15] [16] [17]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: In neuroblastoma, MYCN amplification is associated to a worse prognosis and is a criteria used in the clinic to provide intensive treatments to children even with localized disease. In correlation with MYCN amplification, upregulation of TWIST1, a transcription factor playing a crucial role in inhibition of apoptosis and differentiation was previously reported. Clinical data set analysis of MYCN, MYC and TWIST1 expression permit us to confirm that TWIST1 expression is upregulated in MYCN amplified neuroblastoma but also in a subset of neuroblastoma harboring high expression of MYCN or MYC without gene amplification. In silico analyses reveal the presence of several MYC regulatory motifs (E-Boxes and INR) within the TWIST1 promoter. Using gel shift assay and reporter activity assays, we demonstrate that both N-Myc and c-Myc proteins can bind and activate the TWIST1 promoter. Therefore, we propose TWIST1 as a direct MYC transcriptional target. Copyright © 2014. Published by Elsevier Ireland Ltd.
    Cancer Letters 12/2014; 357(1). DOI:10.1016/j.canlet.2014.11.056 · 5.62 Impact Factor
  • Source
    • "Chromatin samples were prepared as previously described [45]. Neurospheres were crosslinked as monolayers. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Here we examine the functions of the Myc cofactor and histone acetyltransferase, GCN5/KAT2A, in neural stem and precursor cells (NSC) using a conditional knockout approach driven by nestin-cre. Mice with GCN5-deficient NSC exhibit a 25% reduction in brain mass with a microcephaly phenotype similar to that observed in nestin-cre driven knockouts of c- or N-myc. In addition, the loss of GCN5 inhibits precursor cell proliferation and reduces their populations in vivo, as does loss of N-myc. Gene expression analysis indicates that about one-sixth of genes whose expression is affected by loss of GCN5 are also affected in the same manner by loss of N-myc. These findings strongly support the notion that GCN5 protein is a key N-Myc transcriptional cofactor in NSC, but are also consistent with recruitment of GCN5 by other transcription factors and the use by N-Myc of other histone acetyltransferases. Putative N-Myc/GCN5 coregulated transcriptional pathways include cell metabolism, cell cycle, chromatin, and neuron projection morphogenesis genes. GCN5 is also required for maintenance of histone acetylation both at its putative specific target genes and at Myc targets. Thus, we have defined an important role for GCN5 in NSC and provided evidence that GCN5 is an important Myc transcriptional cofactor in vivo.
    PLoS ONE 06/2012; 7(6):e39456. DOI:10.1371/journal.pone.0039456 · 3.23 Impact Factor
  • Source
    • "Data suggests that during iPS cell formation, Myc represses differentiation-associated genes [28] and may not have a key role in directly maintaining expression of pluripotency factors. However, in neuroblastoma and other tumors, some pluripotency genes such as lif, lin28b, Klf2, and Klf4 are N-Myc targets for activation, while a subset of these genes is also regulated in NSC by N-Myc [29]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Separate murine knockout (KO) of either c- or N-myc genes in neural stem and precursor cells (NSC) driven by nestin-cre causes microcephaly. The cerebellum is particularly affected in the N-myc KO, leading to a strong reduction in cerebellar granule neural progenitors (CGNP) and mature granule neurons. In humans, mutation of N-myc also causes microcephaly in Feingold Syndrome. We created a double KO (DKO) of c- and N-myc using nestin-cre, which strongly impairs brain growth, particularly that of the cerebellum. Granule neurons were almost absent from the Myc DKO cerebellum, and other cell types were relatively overrepresented, including astroglia, oligodendrocytes, and Purkinje neurons. These findings are indicative of a profound disruption of cell fate of cerebellar stem and precursors. DKO Purkinje neurons were strikingly lacking in normal arborization. Inhibitory neurons were ectopic and exhibited very abnormal GAD67 staining patterns. Also consistent with altered cell fate, the adult DKO cerebellum still retained a residual external germinal layer (EGL). CGNP in the DKO EGL were almost uniformly NeuN and p27KIP1 positive as well as negative for Math1 and BrdU at the peak of normal cerebellar proliferation at P6. The presence of some mitotic CGNP in the absence of S phase cells suggests a possible arrest in M phase. CGNP and NSC metabolism also was affected by loss of Myc as DKO cells exhibited weak nucleolin staining. Together these findings indicate that c- and N-Myc direct cerebellar development by maintaining CGNP and NSC populations through inhibiting differentiation as well as directing rapid cell cycling and active cellular metabolism. Electronic supplementary material The online version of this article (doi:10.1007/s12311-010-0190-9) contains supplementary material, which is available to authorized users.
    The Cerebellum 12/2010; 9(4):537-47. DOI:10.1007/s12311-010-0190-9 · 2.72 Impact Factor
Show more

Preview (3 Sources)

14 Reads
Available from