346 EXPRESSION OF IMPRINTED NONCODING RNA FROM THE DLK1-DIO3 LOCUS IN HUMAN EMBRYONIC STEM CELLS ADVANTAGES NEURAL LINEAGE DIFFERENTIATION
C.-F. Mo A , F.-C. Wu B , K.-Y. T. Tai C , W.-C. Chang A , K.-W. Chang C , H.-C. Kuo D , H.-N. Ho B , H.-F. Chen B and S.-P. Lin A EA Institute of Biotechnology, National Taiwan University, Taipei, Taiwan;
B Department of Obstetrics & Gynecology, National Taiwan University Hospital, Taipei, Taiwan;
C Genome and Systems Biology Degree Program, Academia Sinica, Taipei, Taiwan;
D Genomic Research Center, Academia Sinica, Taipei, Taiwan;
E Agricultural Biotechnology Research Centre, Academia Sinica, Taipei, Taiwan
Reproduction, Fertility and Development 27(1) 261-261 https://doi.org/10.1071/RDv27n1Ab346
Published: 4 December 2014
Abstract
Pluripotent stem cells are increasingly used for therapeutic models, including transplantation of neural progenitors derived from human embryonic stem cells (hESC). Recently, long noncoding RNA (lncRNA), including Maternally Expressed Gene 3 (MEG3) derived from the DLK1-DIO3-imprinted locus, were found to be expressed during neural developmental events. Their deregulations are associated with various neurological diseases. The DLK1-DIO3-imprinted locus encodes abundant noncoding RNA (ncRNA) that are regulated by differential methylation on the locus. The aim of our research was to study the correlation between the DLK1-DIO3-derived ncRNA and the capacity of hESC neural lineage differentiation. We classified hESC into MEG3-ON and MEG3-OFF based on the expression levels of MEG3 as well as its downstream microRNA by qRT-PCR. Initial embryoid body (EB) formation was conducted to examine the 3 germ layer's differentiation ability. Complementary DNA microarray was used to analyse the gene expression profiles of hESC. Directed neural lineage differentiation was performed, followed by analysis of neural lineage marker expression levels and neurite formation via qRT-PCR and immunocytochemistry methods to investigate the capacity of neural differentiation in MEG3-ON and MEG3-OFF hESC. As for statistics, error bars indicate standard error of the mean. Student's t-test was used for calculating P-values, and a P-value of less than 0.05 was considered to be significant. Our results showed that MEG3-ON and MEG3-OFF hESC differed greatly in DLK1-DIO3-derived ncRNA expression levels, but had comparable pluripotency gene expression profiles. Genes related to nervous system development and neural cancers were differentially expressed in MEG3-OFF hESC, where DLK1-DIO3-derived ncRNA were repressed compared to MEG3-ON ones before differentiation. In neural lineage-like cells derived from MEG3-OFF hESC, lower expression levels of neural lineage markers and impaired neurite formation were observed compared to MEG3-ON hESC at the same time points after differentiation. We suggest that the expression of DLK1-DIO3-derived lncRNA, MEG3, can be used as a simple and effective screening criterion for identifying MEG3-ON hESC with activation of DLK1-DIO3-imprinted ncRNA as starting materials to benefit neural lineage-associated studies.
