PRDM14 suppresses expression of differentiation marker genes in human embryonic stem cells

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Abstract

PRDM14 was identified by microarray analysis and was expressed in specifically undifferentiated human ES cells. PRDM14 protein is thought to regulate gene transcription in human ES cells, as it contains a PR domain, a subtype of the SET domain which catalyzes histone methylation. To analyze the function of PRDM14, we performed knock-down and forced expression of PRDM14 in human ES cells. Knock-down of PRDM14 by siRNA induced expression of early differentiation marker genes. Forced expression of PRDM14 suppressed expression of differentiation marker genes in the embryoid body. These results suggest that PRDM14 is involved in the maintenance of the self-renewal of human ES cells by suppression of gene expression.

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Materials and Methods

Cell culture. The human ES cell lines, KhES-1, KhES-2, and KhES-3, were maintained in human ES cell medium on a feeder layer of mouse embryonic fibroblasts (MEF) inactivated with mitomycin C (Wako Pure Chemical Industries, Ltd., Osaka, Japan) as previously described [16], [23]. The MEF-conditioned medium (CM) was prepared as previously described [16].

For feeder-free culture, human ES cells were dissociated into small clumps and plated on dishes coated with Matrigel (Becton, Dickinson and

Identification of genes specifically expressed in undifferentiated human ES cells

Genes that are specifically expressed in undifferentiated human ES cells have the potential to play important roles in human ES cell self-renewal. To find genes that can elucidate the mechanism of human ES cell self-renewal, we conducted microarray analysis (data not shown) using the total RNA from KhES-1 cells (a human ES cell line) and TIG-3 cells (a fetal lung fibroblast cell line, as differentiated cells), and identified 89 genes whose mRNA levels are upregulated in KhES-1 cells compared to

Discussion

In the present study, we performed a microarray analysis to find genes that might be involved in maintenance of human ES cell self-renewal, and identified 89 candidate genes whose expressions are upregulated in human ES cells compared to TIG-3 cells (Fig. S1 and Table S1). Since many of the genes that play important roles in maintenance of ES cell self-renewal, such as NANOG, POU5F1, SOX2, TDGF1, LEFTY1, LEFTY2, ZIC3, and SALL4[4], [5], [6], [7], [8], [9], [10], [27], [28], [29], are

Acknowledgments

We thank Takahiro Tougan of Osaka University and the members of the laboratories of N. Nakatsuji and H. Suemori for valuable discussions and technical assistance. This work was supported in part by the National BioResource Project of the Ministry of Education, Culture, Sports, Science, and Technology (MEXT) of Japan, the Japan Society for the Promotion of Science, and by New Energy and Industrial Technology Development Organization (NEDO) of Japan.

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