Loss of non-coding RNA expression from the DLK1-DIO3 imprinted locus correlates with reduced neural differentiation potential in human embryonic stem cell lines

Chu Fan Mo, Fang Chun Wu, Kang Yu Tai, Wei Chun Chang, Kai Wei Chang, Hung Chih Kuo, Hong Nerng Ho, Hsin Fu Chen, Shau Ping Lin

研究成果: 雜誌貢獻文章同行評審

42 引文 斯高帕斯(Scopus)

摘要

Introduction: Pluripotent stem cells are increasingly used to build therapeutic models, including the transplantation of neural progenitors derived from human embryonic stem cells (hESCs). Recently, long non-coding RNAs (lncRNAs), including delta-like homolog 1 gene and the type III iodothyronine deiodinase gene (DLK1-DIO3) imprinted locus-derived maternally expressed gene 3 (MEG3), were found to be expressed during neural development. The deregulation of these lncRNAs is associated with various neurological diseases. The imprinted locus DLK1-DIO3 encodes abundant non-coding RNAs (ncRNAs) that are regulated by differential methylation of the locus. We aim to study the correlation between the DLK1-DIO3-derived ncRNAs and the capacity of hESCs to differentiate into neural lineages. Methods: We classified hESC sublines into MEG3-ON and MEG3-OFF based on the expression levels of MEG3 and its downstream microRNAs as detected by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). A cDNA microarray was used to analyze the gene expression profiles of hESCs. To investigate the capacity of neural differentiation in MEG3-ON and MEG3-OFF hESCs, we performed neural lineage differentiation followed by neural lineage marker expression and neurite formation analyses via qRT-PCR and immunocytochemistry, respectively. MEG3-knockdown via small interfering RNA (siRNA) and small hairpin RNA (shRNA) was used to investigate the potential causative effect of MEG3 in regulating neural lineage-related gene expression. Results: DLK1-DIO3-derived ncRNAs were repressed in MEG3-OFF hESCs compared with those in the MEG3-ON hESCs. The transcriptome profile indicated that many genes related to nervous system development and neural-type tumors were differentially expressed in MEG3-OFF hESCs. Three independent MEG3-knockdown assays using different siRNA and shRNA constructs consistently resulted in downregulation of some neural lineage genes. Lower expression levels of stage-specific neural lineage markers and reduced neurite formation were observed in neural lineage-like cells derived from MEG3-OFF-associated hESCs compared with those in the MEG3-ON groups at the same time points after differentiation. Conclusions: Repression of ncRNAs derived from the DLK1-DIO3 imprinted locus is associated with reduced neural lineage differentiation potential in hESCs.
原文英語
文章編號1
期刊Stem Cell Research and Therapy
6
發行號1
DOIs
出版狀態已發佈 - 1月 5 2015
對外發佈

ASJC Scopus subject areas

  • 醫藥(雜項)
  • 分子醫學
  • 生物化學、遺傳與分子生物學(雜項)
  • 細胞生物學

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