title: Heterogeneous lineage marker expression in naive embryonic stem cells is mostly due to spontaneous differentiation.
abstract: Populations of cultured mouse embryonic stem cells (ESCs) exhibit a subfraction of cells expressing uncharacteristically low levels of pluripotency markers such as Nanog. Yet, the extent to which individual Nanog-negative cells are differentiated, both from ESCs and from each other, remains unclear. Here, we show the transcriptome of Nanog-negative cells exhibits expression of classes of genes associated with differentiation that are not yet active in cells exposed to differentiation conditions for one day. Long non-coding RNAs, however, exhibit more changes in expression in the one-day-differentiated cells than in Nanog-negative cells. These results are consistent with the concept that Nanog-negative cells may contain subpopulations of both lineage-primed and differentiated cells. Single cell analysis showed that Nanog-negative cells display substantial and coherent heterogeneity in lineage marker expression in progressively nested subsets of cells exhibiting low levels of Nanog, then low levels of Oct4, and then a set of lineage markers, which express intensely in a small subset of these more differentiated cells. Our results suggest that the observed enrichment of lineage-specific marker gene expression in Nanog-negative cells is associated with spontaneous differentiation of a subset of these cells rather than the more random expression that may be associated with reversible lineage priming.
authors: Department of Bioengineering, University of Pennsylvania, Philadelphia, PA 19104, USA.
authors: Gautham
authors: G
authors: Nair
authors: Instituto de Medicina Molecular and Instituto de Histologia e Biologia do Desenvolvimento, Faculdade de Medicina da Universidade de Lisboa, Lisboa, Portugal.|Champalimaud Neuroscience Programme, Champalimaud Centre for the Unknown, Doca de Pedroucos, Lisboa, Portugal.
authors: Elsa
authors: E
authors: Abranches
authors: Instituto de Medicina Molecular and Instituto de Histologia e Biologia do Desenvolvimento, Faculdade de Medicina da Universidade de Lisboa, Lisboa, Portugal.|Champalimaud Neuroscience Programme, Champalimaud Centre for the Unknown, Doca de Pedroucos, Lisboa, Portugal.
authors: Ana M V
authors: AM
authors: Guedes
authors: Instituto de Medicina Molecular and Instituto de Histologia e Biologia do Desenvolvimento, Faculdade de Medicina da Universidade de Lisboa, Lisboa, Portugal.|Champalimaud Neuroscience Programme, Champalimaud Centre for the Unknown, Doca de Pedroucos, Lisboa, Portugal.
authors: Domingos
authors: D
authors: Henrique
authors: Department of Bioengineering, University of Pennsylvania, Philadelphia, PA 19104, USA.
authors: Arjun
authors: A
authors: Raj
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism*; D002454:Cell Differentiation / Q000235:genetics; D002460:Cell Line; D019070:Cell Lineage / Q000235:genetics; D020869:Gene Expression Profiling; D016678:Genome; D018398:Homeodomain Proteins / Q000378:metabolism; D017404:In Situ Hybridization, Fluorescence; D051379:Mice; D000066450:Mouse Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism*; D000071317:Nanog Homeobox Protein; D011485:Protein Binding; D062085:RNA, Long Noncoding / Q000235:genetics / Q000378:metabolism; D059010:Single-Cell Analysis; D014157:Transcription Factors / Q000378:metabolism; D059467:Transcriptome / Q000235:genetics
chemical_list: D015415:Biomarkers; D018398:Homeodomain Proteins; D000071317:Nanog Homeobox Protein; C475540:Nanog protein, mouse; D062085:RNA, Long Noncoding; D014157:Transcription Factors