Dissecting Oct3/4-Regulated Gene Networks in Embryonic Stem Cells by Expression Profiling

Abstract

POU transcription factor Pou5f1 (Oct3/4) is required to maintain ES cells in an undifferentiated state. Here we show that global expression profiling of Oct3/4-manipulated ES cells delineates the downstream target genes of Oct3/4. Combined with data from genome-wide chromatin-immunoprecipitation (ChIP) assays, this analysis identifies not only primary downstream targets of Oct3/4, but also secondary or tertiary targets. Furthermore, the analysis also reveals that downstream target genes are regulated either positively or negatively by Oct3/4. Identification of a group of genes that show both activation and repression depending on Oct3/4 expression levels provides a possible mechanism for the requirement of appropriate Oct3/4 expression to maintain undifferentiated ES cells. As a proof-of-principle study, one of the downstream genes, Tcl1, has been analyzed in detail. We show that Oct3/4 binds to the promoter region of Tcl1 and activates its transcription. We also show that Tcl1 is involved in the regulation of proliferation, but not differentiation, in ES cells. These findings suggest that the global expression profiling of gene-manipulated ES cells can help to delineate the structure and dynamics of gene regulatory networks.

The Tcl1 oncogene defines secondary hair germ cells differentiation at catagen–telogen transition and affects stem-cell marker CD34 expression

“…However, it is interesting to observe that in the epidermis, deletion of Akt1 causes thinner epidermal layers and retarded-HF development, whereas double mutants (Akt1 À/À Akt2 À/À and Akt1 À/À Akt3 À/À ) (Peng et al, 2003;Yang et al, 2005) show more severe defects, and transgenic mice expressing Akt-Mer fusion proteins induces epidermal hyperplasia and proliferation of epidermal progenitors leading to epidermal and follicular hyperplasia (Murayama et al, 2007). As already pointed out before, Tcl1 is involved in the normal-self renewal of ES cells (Glover et al, 2006;Matoba et al, 2006;GalanCaritad et al, 2007) and upstream genes such as transcription factor Oct4, which directly activates Tcl1 in ES cells and is expressed in human HFs (Yu et al, 2006), should also be taken into account.…”

“…If the expression of Tcl1 is maintained constantly within the same cells, as it seems to be in other tissues such as B cells and embryo, it would implicate that proliferating cells that derive from HG-cells, during catagen-telogen transition, are those starting the new HF cycle and repopulating the bulge ( Figure 5). As in the embryo, where Tcl1 is necessary in the first divisions of proliferating cells of the preimplantation embryo, and for self-renewal but not differentiation of their derived ES-cells (Matoba et al, 2006;Galan-Caritad et al, 2007) in the HF, Tcl1 is important very early during the anagen proliferation of secondary HG-cells, as beyond the third day after depilation the expression of Tcl1 is no more detectable.…”

“…In particular, it plays an important role in the latter as, in Tcl1 À/À mice, preimplantation embryos are not able to progress beyond the 4-cell stage, in vitro (Narducci et al, 2002). In ES cells, Tcl1 acts in proliferation and normal self-renewal under the direct activation of OCT3/4, Zfx and KLF5 transcription factors (Matoba et al, 2006;Galan-Caritad et al, 2007;Ema et al, 2008). Therefore, Tcl1 is not ubiquitously expressed, but seems to be tightly regulated in cells that undergo different waves of proliferative signals to complete their differentiation programs: pro-B cells in the bone marrow, the mantle lymphocytes of peripheral lymph nodes in which B cells must encounter the antigen and go through new waves of mitosis, the fertilized egg before zygotic gene activation when the cell, still under maternal control, must divide rapidly after having been blocked in metaphase II for a long time and ES cells.…”

The BRG1 ATPase of chromatin remodeling complexes is involved in modulation of mesenchymal stem cell senescence through RB–P53 pathways

“…For a third group of genes, we obtained reproducible reverse transcriptase-PCR evaluation ( Figure 3). In particular, we detected the expression of OCT3, NANOG and KLF4, which are part of the core transcriptional circuitry for regulation of embryonic stem cell properties (Matoba et al, 2006;Masui et al, 2007). Of great interest is that BRG1 downregulation completely abolished NANOG expression ( Figure 3).…”

Dynamic expression of Thyrotropin‐releasing hormone in the mouse definitive endoderm

“…To confirm the SAGE data, we characterized Trh expression using WISH where we see expression at E6.5 but not at E5.5 (data not shown). Of interest, it has recently been shown that Trh may be a direct transcriptional target of POU5F1, a well-characterized marker of pluripotency in embryonic stem cells and the epiblast (Loh et al, 2006;Matoba et al, 2006). Pou5f1 expression in the epiblast overlaps with that of Trh, suggesting that POU5F1 may in part be regulating Trh expression at this stage (Rosner et al, 1990).…”