MicroRNA-125b Promotes Neuronal Differentiation in Human Cells by Repressing Multiple Targets

Whitehead Institute for Biomedical Research, 9 Cambridge Center, Suite 601, Cambridge, MA 02142, USA.
Molecular and Cellular Biology (Impact Factor: 4.78). 08/2009; 29(19):5290-305. DOI: 10.1128/MCB.01694-08
Source: PubMed


MicroRNAs (miRNAs) are a class of small noncoding RNAs that regulate gene expression at the posttranscriptional level. Research on miRNAs has highlighted their importance in neural development, but the specific functions of neurally enriched miRNAs remain poorly understood. We report here the expression profile of miRNAs during neuronal differentiation in the human neuroblastoma cell line SH-SY5Y. Six miRNAs were significantly upregulated during differentiation induced by all-trans-retinoic acid and brain-derived neurotrophic factor. We demonstrated that the ectopic expression of either miR-124a or miR-125b increases the percentage of differentiated SH-SY5Y cells with neurite outgrowth. Subsequently, we focused our functional analysis on miR-125b and demonstrated the important role of this miRNA in both the spontaneous and induced differentiations of SH-SH5Y cells. miR-125b is also upregulated during the differentiation of human neural progenitor ReNcell VM cells, and miR-125b ectopic expression significantly promotes the neurite outgrowth of these cells. To identify the targets of miR-125b regulation, we profiled the global changes in gene expression following miR-125b ectopic expression in SH-SY5Y cells. miR-125b represses 164 genes that contain the seed match sequence of the miRNA and/or that are predicted to be direct targets of miR-125b by conventional methods. Pathway analysis suggests that a subset of miR-125b-repressed targets antagonizes neuronal genes in several neurogenic pathways, thereby mediating the positive effect of miR-125b on neuronal differentiation. We have further validated the binding of miR-125b to the miRNA response elements of 10 selected mRNA targets. Together, we report here for the first time the important role of miR-125b in human neuronal differentiation.

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Available from: Beiyan Zhou,
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    • "We asked whether LSD1+8a expression was also dynamically regulated during human neuronal differentiation using SH-SY5Y cells, which are human neuronal cells derived from neuroblastoma. Sequential exposure of these cells to retinoic acid (RA) for 4 days (day D0 to D4) and then to brainderived neurotrophic factor (BDNF) in a serum-free medium for 5 days (day D4/B0 to B5) yields a homogeneous population of differentiated neuronal cells that possess many features of primary neurons (Figure S1A) (Encinas et al., 2000; Le et al., 2009). While exposure of SH-SY5Y cells to RA slightly reduced the level of both types of LSD1 transcripts (LSD1-8a and LSD1+8a), BDNF treatment increased the expression level of LSD1+8a isoforms more than that of LSD1-8a isoforms between day B0 and B5 (3.85 ± 0.3 versus 1.47 ± 0.15) (Figure 1A). "
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    ABSTRACT: Lysine-specific demethylase 1 (LSD1) has been reported to repress and activate transcription by mediating histone H3K4me1/2 and H3K9me1/2 demethylation, respectively. The molecular mechanism that underlies this dual substrate specificity has remained unknown. Here we report that an isoform of LSD1, LSD1+8a, does not have the intrinsic capability to demethylate H3K4me2. Instead, LSD1+8a mediates H3K9me2 demethylation in collaboration with supervillin (SVIL), a new LSD1+8a interacting protein. LSD1+8a knockdown increases H3K9me2, but not H3K4me2, levels at its target promoters and compromises neuronal differentiation. Importantly, SVIL co-localizes to LSD1+8a-bound promoters, and its knockdown mimics the impact of LSD1+8a loss, supporting SVIL as a cofactor for LSD1+8a in neuronal cells. These findings provide insight into mechanisms by which LSD1 mediates H3K9me demethylation and highlight alternative splicing as a means by which LSD1 acquires selective substrate specificities (H3K9 versus H3K4) to differentially control specific gene expression programs in neurons. Copyright © 2015 Elsevier Inc. All rights reserved.
    Molecular cell 02/2015; 57(6). DOI:10.1016/j.molcel.2015.01.010 · 14.02 Impact Factor
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    • "These experiments showed that overexpression of LSD1 inhibited neurite extension and the expression of neuronal markers in differentiating SH- SY5Y cells, effects that could be abolished by co-overexpression of Jade-2 but not Jade-2-C202A or Jade-2-C243A (Figures 7C and 7D). Full differentiation of SH-SY5Y cells by sequential exposure to RA and brain-derived neurotrophic factor (BDNF0) (Encinas et al., 2000; Le et al., 2009) (Figure S5B) showed that both mRNA and protein levels of LSD1 decreased, whereas the expression of Jade-2 did not change during neuronal differentiation (Figure S5C). Subsequently, we showed that LSD1 inhibits neuronal differentiation of SH-SY5Y cells, whereas Jade-2 promotes this process, which could be offset by simultaneous overexpression of LSD1 (Figures S5D and S5E). "
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    ABSTRACT: Histone H3K4 demethylase LSD1 plays an important role in stem cell biology, especially in the maintenance of the silencing of differentiation genes. However, how the function of LSD1 is regulated and the differentiation genes are derepressed are not understood. Here, we report that elimination of LSD1 promotes embryonic stem cell (ESC) differentiation toward neural lineage. We showed that the destabilization of LSD1 occurs posttranscriptionally via the ubiquitin-proteasome pathway by an E3 ubiquitin ligase, Jade-2. We demonstrated that Jade-2 is a major LSD1 negative regulator during neurogenesis in vitro and in vivo in both mouse developing cerebral cortices and zebra fish embryos. Apparently, Jade-2-mediated degradation of LSD1 acts as an antibraking system and serves as a quick adaptive mechanism for re-establishing epigenetic landscape without more laborious transcriptional regulations. As a potential anticancer strategy, Jade-2-mediated LSD1 degradation could potentially be used in neuroblastoma cells to induce differentiation toward postmitotic neurons.
    Molecular Cell 07/2014; 55(3). DOI:10.1016/j.molcel.2014.06.006 · 14.02 Impact Factor
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    • "Protein levels and function of p53 have been shown to be regulated by miRNAs including miR-125 [43], miR-18 [23], and miR-504 [28]. The miR-504 has been reported as a negative regulator of p53 through its direct binding in the p53 3' untranslated region, thereby decreasing p53 function and protein expression, without affecting mRNA levels [28]. "
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    ABSTRACT: The expression of TFF1 is frequently down-regulated in human gastric cancer whereas its knockout leads to the development of gastric adenomas and carcinomas in mouse models. The molecular mechanisms underlying the TFF1 tumor suppressor functions remain unclear. In this study, we demonstrate, using colony formation assay and Annexin V staining, that reconstitution of TFF1 expression in gastric cancer cell models suppresses cell growth and promotes cell death. Furthermore, using a tumor xenograft mouse model of gastric cancer, we demonstrated that reconstitution of TFF1 suppresses tumor growth in vivo. The results from PG13-luciferase reporter assay and Western blot analysis indicated that TFF1 promotes the expression and transcription activity of p53 protein. Further analysis using cycloheximide-based protein assay and quantitative real-time PCR data suggested that TFF1 does not interfere with p53 mRNA levels or protein stability. Alternatively, we found that the reconstitution of TFF1 down-regulates miR-504, a negative regulator of p53. Western blot analysis data demonstrated that miR-504 abrogates TFF1-induced p53 protein expression and activity. In conclusion, the in vitro and in vivo data demonstrate, for the first time, a novel mechanism by which the tumor suppressor functions of TFF1 involve activation of p53 through down-regulation of miR-504.
    Oncotarget 07/2014; 5(14). DOI:10.18632/oncotarget.2156 · 6.36 Impact Factor
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