Estrogen-Related Receptor Beta Interacts with Oct4 To Positively Regulate Nanog Gene Expression

Abstract

Embryonic stem (ES) cell self-renewal is regulated by transcription factors, including Oct4, Sox2, and Nanog. A number of additional transcriptional regulators of ES cell self-renewal have recently been identified, including the orphan nuclear receptor estrogen-related receptor beta (Esrrb). However, the mode of action of Esrrb in ES cells is unknown. Here, using an Oct4 affinity screen, we identify Esrrb as an Oct4 partner protein. Esrrb can interact with Oct4 independently of DNA. Esrrb is recruited near the Oct-Sox element in the Nanog proximal promoter, where it positively regulates Nanog expression. Esrrb recruitment to the Nanog promoter requires both the presence of Oct4 and a degenerate estrogen-related receptor DNA element. Consistent with its role in Nanog regulation, expression of the Esrrb protein within the Oct4-positive ES cell population is mosaic and correlates with the mosaic expression of the Nanog protein. Together with previous reports that Nanog may regulate Esrrb gene expression, our results suggest that Esrrb and Nanog act as part of a feedback regulatory circuit that modulates the fluctuating self-renewal capacity of ES cell populations.The self-renewal of mouse ES cells is regulated by a network of transcription factors that includes Oct4, Nanog, and Sox2 (22). The expression level of Oct4 protein needs to be kept within a tight range in order to maintain ES cell self-renewal (23). Decreasing Oct4 levels below 50% induces differentiation into the trophectoderm, whereas a twofold increase causes differentiation into cells expressing markers of the endoderm and mesoderm (23). In contrast, overexpression of Nanog allows mouse embryonic stem (ES) cells to remain undifferentiated in the absence of the otherwise requisite stimulation by leukemia inhibitory factor and bone morphogenetic protein (5,19,34). Oct4 is thought to act together with Sox2 by binding to adjacent cognate DNA sequences in many genes (1), including Nanog (14, 26). Genome-wide chromatin immunoprecipitation (ChIP) studies have suggested that composite Oct-Sox motifs regulate the expression of many genes in mouse and human ES cells (2, 16). Recent evidence has shown that the critical role of Sox2 in maintaining ES cell self-renewal is regulating Oct4 expression, suggesting that the secondary role of gene regulation via Oct-Sox motifs is performed redundantly with Sox4, Sox11, and Sox15 (18).Recent reports have expanded the list of factors that contribute to ES cell self-renewal. Wang et al. (30) reported a proteomic analysis of interactors of Oct4 and Nanog and suggested that some Nanog interactors may assist in Nanogmediated gene regulation. A separate study using an RNA interference (RNAi) screen found that depletion of estrogenrelated receptor beta (Esrrb), Tbx3, or Tcl1 resulted in ES cell differentiation but that this differentiation could be attenuated by overexpression of Nanog (13). However, it is unclear how any of these novel regulatory factors mediate their function. Here we use an unbiased analysis of Oct4 binding...

Zfp281 mediates Nanog autorepression through recruitment of the NuRD complex and inhibits somatic cell reprogramming

“…3B), we speculate that the activation function of ectopic Nanog observed in Zfp281 −/− ESCs is more likely resulted from transactivation by other pluripotency factors that act on the endogenous Nanog locus. For example, Oct4, Esrrb, and Zfp143 are known to form heterodimers and directly transactivate Nanog promoter activity (23)(24)(25), and Tbx3 was shown to predominantly stimulate Nanog expression in maintaining pluripotency (26). We confirmed Doxdependent up-regulation of Oct4, Esrrb, Zfp143, Tbx3, and Rex1 in Zfp281 −/− ESCs but only slightly up-regulated or unchanged levels of these genes in Zfp281 +/+ ESCs (Fig.…”

Esrrb extinction triggers dismantling of naïve pluripotency and marks commitment to differentiation

“…Moreover, quantitation of pre‐mRNA shows that Esrrb transcription responds to NANOG relocalization within minutes, indicating that Esrrb is a primary direct NANOG target. Transient transfection assays also indicate that ESRRB can contribute to luciferase reporter expression driven by the Nanog promoter (van den Berg et al , 2008). To further investigate the effect of ESRRB on Nanog gene expression and of NANOG on Esrrb gene expression, immunofluorescence analysis was performed on ESCs overexpressing NANOG or ESRRB.…”

Esrrb directly binds to Gata6 promoter and regulates its expression with Dax1 and Ncoa3

“…Estrogen-related receptor beta (Esrrb) is considered to be a member of the core transcriptional regulatory network in ES cells [17,18], and has been associated with Oct3/4 [13,19], Nanog [10,20], and Dax1 [21], and is reportedly required for self-renewal of ES cells [22,23]. Moreover, Esrrb has been shown to regulate the self-renewal capacity of ES cells downstream of Nanog and Gsk3-Tcf3 pathways [24,25].…”