Opposing Putative Roles for Canonical and Noncanonical NFκB Signaling on the Survival, Proliferation, and Differentiation Potential of Human Embryonic Stem Cells

Institute of Human Genetics, International Centre for Life, Newcastle University, Newcastle Upon Tyne, United Kingdom.
Stem Cells (Impact Factor: 6.52). 11/2010; 28(11):1970-80. DOI: 10.1002/stem.528
Source: PubMed


The canonical and noncanonical NFκB signaling pathways regulate a variety of cellular activities; however, their functions in human embryonic stem cells (hESCs) have not been fully investigated. Expression studies during hESC differentiation indicated a significant increase in the expression of two key components of the canonical NFκB pathway (p50 and Ser529 phosphorylated form of p65) as well as a significant reduction in expression of key components of the noncanonical NFκB pathway [v-rel reticuloendotheliosis viral oncogene homolog B (RELB), p52, NIK]. Inhibition of canonical NFκB resulted in hESC apoptosis, changes in cell cycle distribution, and reduced hESC proliferation. In addition, inhibition of canonical NFκB was associated with significant changes in NANOG and OCT4 expression, suppression of differentiation toward all primitive extraembryonic and embryonic lineages with the exception of primitive ectoderm and ectodermal lineages. Inhibition of noncanonical NFκB via small interfering RNA-mediated downregulation of RELB resulted in reduced hESC proliferation and opposite changes to expression of key differentiation lineage markers genes when compared with downregulation of canonical NF-κB. Chromatin immunoprecipitation assays indicated binding of p65 and RELB to regulatory regions of key differentiation marker genes suggesting a direct transcriptional role for both branches of this pathway in hESC. These findings coupled with opposing trends in expression of key components during hESC differentiation, suggests a fine and opposing balance between the two branches of NFκB signaling pathways and their involvement in two distinct processes: the canonical pathway regulating hESC differentiation and the noncanonical pathway maintaining hESC pluripotency.

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Available from: Stuart P Atkinson, Aug 27, 2014
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    • "Armstrong et al. reported specific inhibition of NF-κB induces prominent differentiation of human ES cells [4], but the same group also reported the activation of NF-κB signaling pathway during differentiation as a result of embryoid body (EB) formation [25]. As Armstrong et al. reported, we confirmed augmented NF-κB activity in undifferentiated human ES cells and down-regulation of NF-κB activity upon monolayer differentiation (Figure S1). "
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    ABSTRACT: NF-κB signaling plays an essential role in maintaining the undifferentiated state of embryonic stem (ES) cells. However, opposing roles of NF-κB have been reported in mouse and human ES cells, and the role of NF-κB in human induced pluripotent stem (iPS) cells has not yet been clarified. Here, we report the role of NF-κB signaling in maintaining the undifferentiated state of human iPS cells. Compared with differentiated cells, undifferentiated human iPS cells showed an augmentation of NF-κB activity. During differentiation induced by the removal of feeder cells and FGF2, we observed a reduction in NF-κB activity, the expression of the undifferentiation markers Oct3/4 and Nanog, and the up-regulation of the differentiated markers WT-1 and Pax-2. The specific knockdown of NF-κB signaling using p65 siRNA also reduced the expression of Oct3/4 and Nanog and up-regulated WT-1 and Pax-2 but did not change the ES-like colony formation. Our results show that the augmentation of NF-κB signaling maintains the undifferentiated state of human iPS and suggest the importance of this signaling pathway in maintenance of human iPS cells.
    PLoS ONE 02/2013; 8(2):e56399. DOI:10.1371/journal.pone.0056399 · 3.23 Impact Factor
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    • "However, several studies have shown that nonclassical pathway plays important role in the differentiation of ES cells and other tissue stem cells. For example, the canonical NFκB pathway regulates differentiation while the noncanonical pathway maintains the pluripotency of human ES cells (Yang et al., 2010). The p100/RelB signaling positively and intrinsically regulates self-renewal of hematopoietic stem/progenitor cell (Zhao et al., 2012). "
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    05/2012; 7(4):277-291. DOI:10.1007/s11515-012-1233-z
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    ABSTRACT: NF-kappaB is involved in many biological processes including proliferation, survival, and differentiation. Because human embryonic stem (ES) cells have the potential to differentiate to various lineages, understanding mechanisms involved in stemness and lineage differentiation is an important issue. We investigated expression of NF-kappaB in the human ES cell lines SNUhES3 and MizhES4 and found that expression of NF-kappaB mRNA and protein in these two cell lines was significantly lower compared to those of other adult cell lines. However, when SNUhES3 cells were induced to differentiate by retinoic acid, expression levels of NF-kappaB significantly increased compared to undifferentiated SNUhES3 cells. As the components of tumor necrosis factor-alpha (TNF-alpha) signaling are expressed comparably in undifferentiated and differentiated SNUhES3 cells, we examined the responsiveness of SNUhES3 cells to treatment with TNF-alpha, an agonist of NF-kappaB signaling. Nuclear localization of NF-kappaB in response to TNF-alpha was evident in differentiated, but not undifferentiated, SNUhES3 cells. In agreement with this observation, induction of interleukin-8 (IL-8) in response to TNF-alpha was seen only in differentiated SNUhES3 cells. On the basis of an IkappaB kinase (IKK) inhibitor study, expression of IL-8 induced by TNF-alpha was dependent on NF-kappaB activity. Taken together, our results suggest that expression and activity of NF-kappaB is comparatively low in undifferentiated human ES cells, but increases during differentiation of the ES cells.
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