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Title: MicroRNA-29 Impairs the Early Phase of Reprogramming Process by Targeting Active DNA Demethylation and Wnt Signaling      
keywords:
Transcriptome or Gene expression
ID:
PRJNA345375
description:
Somatic cell reprogramming by transcription factors and other modifiers such as small molecules or miRNAs has opened broad avenues for the study of developmental processes, cell fate determination, and interplay of molecular mechanisms in signaling pathways. However, many of the mechanisms that drive nuclear reprogramming itself remain yet to be elucidated. Here, we analyzed the role of miR-29 during reprogramming in more detail. Therefore, we evaluated miR-29 expression during reprogramming of fibroblasts transduced with lentiviral OKS and OKSM vectors and we show that addition of c-MYC to the reprogramming factor cocktail decreases miR-29 expression levels. Moreover, we found that transfection of pre-miR-29a strongly decreased OKS-induced formation of GFP+- colonies in MEF-cells from Oct4-eGFP reporter mouse, whereas anti-miR-29a showed the opposite effect. Furthermore, we studied two pathways which are important for reprogramming and which involve miR-29 targets: active DNA-demethylation and Wntsignaling. We show that inhibition of active DNA-demethylation as well as activation of Wnt-signaling leads to decreased reprogramming efficiency. Moreover, transfection of premiR-29 resulted in elevated expression of β-Catenin transcriptional target sFRP2 and increased TCF/LEF-reporter activity. Finally, we report that Gsk3-β is a direct target of miR-29 in MEF-cells. Together, our findings contribute to the understanding of the molecular mechanisms by which miR29 influences reprogramming. Overall design: In order to identify potential miR-29a targets, human BJ1 fibroblasts were transfected with 50 nM of synthetic miR-29a mimics (n=3) or control miR-Ctr molecules (n=3) in 12-well plates, using Lipofectamine RNAiMax (Invitrogen), according to the manufacturer's instructions. Total RNA was extracted 72 hours post-transfection and analyzed.
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landingpage: http://www.ncbi.nlm.nih.gov/bioproject/PRJNA345375
authentication:
none
authorization:
none
ID:
pmid:28038351
name:
Homo sapiens
ncbiID:
ncbitax:9606
abbreviation:
NCBI
homePage: http://www.ncbi.nlm.nih.gov
ID:
SCR:006472
name:
National Center for Biotechnology Information
homePage: http://www.ncbi.nlm.nih.gov/bioproject
ID:
SCR:004801
name:
NCBI BioProject