Differential expression of Nanog 3UTR and coding region mRNA components defines an embryonic stem cell niche

Abstract

SummaryThe known molecular heterogeneity among pluripotent embryonic stem cells (ESCs) in culture may be maladaptive or a beneficial determinant of normal biology. In other systems such as hair and intestine, normal and dynamic heterogeneity among stem cells is necessary for balancing stem cell division and quiescence in order to maintain the stem cell pool. In ESCs, Sox2, Oct4 and Nanog (the SON genes) critically regulate pluripotency but their role in stem cell heterogeneity or pool maintenance is not clear. We previously reported an additional level of gene expression, the unexpected differential expression of mRNA coding region (CDS) and cognate 3′untranslated region (3UTR) components, often revealing novel developmental patterns. Here we show dramatic, patterned, concentric, differential expression of pluripotent gene 3UTR and CDS mRNA components in both undifferentiated ESCs in culture and in developing human and mouse embryos. This differential expression reveals robust pluripotent ESC heterogeneity that provides the underpinnings of a previously unrecognized ESC cell niche structure. A hallmark of the niche is the presence of cells expressing high levels of Nanog 3UTR but only low or no Nanog CDS at structure perimeters, We used a novel bioinformatic approach combining the 3UTR to CDS ratio and gene expression, and identified unique ESC clusters. High Nanog 3UTR cells have lower cell cycle activity and higher quiescence than other ESCs and express a distinct complement of cellular histone marks. These quiescent perimeter cells exchange with interior domains, opening the possibility that they act as cell replenishment reservoirs in the niche or the embryo. Functionally, the Nanog 3UTR, independent of its CDS, is critical, as its selective deletion decreases ESC cell division while increasing trophectoderm markers. Lastly, self-organization of ESCs according to pluripotent gene differential 3UTR and CDS expression is modified by activin signaling, indicating extrinsic modulation of the niche structure. Pluripotent embryonic stem cells, actively display regulated, dynamic differential expression of SON gene mRNA components, allowing the formation of a heterogenous, ordered, stem cell niche bordered by quiescent hub stem cells, thus illuminating the role of SON genes in pluripotent embryonic stem cell pool maintenance. More generally this suggests that differential mRNA component expression may widely define cell types that exist in non-permanent, different states in either development or disease.