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ABSTRACT: The molecular events leading to human embryonic stem cell (hESC) differentiation are the subject of considerable scrutiny. Here, we characterize an in vitro model that permits analysis of the earliest steps in the transition of hESC colonies to squamous epithelium on basic fibroblast growth factor withdrawal. A set of markers (GSC, CK18, Gata4, Eomes, and Sox17) point to a mesendodermal nature of the epithelial cells with subsequent commitment to definitive endoderm (Sox17, Cdx2, nestin, and Islet1). We assayed alterations in the transcriptome in parallel with the distribution of immunohistochemical markers. Our results indicate that the alterations of tight junctions in pluripotent culture precede the beginning of differentiation. We defined this cell population as "specified," as it is committed toward differentiation. The transitional zone between "specified" pluripotent and differentiated cells displays significant up-regulation of keratin-18 (CK18) along with a decrease in the functional activity of gap junctions and the down-regulation of 2 gap junction proteins, connexin 43 (Cx43) and connexin 45 (Cx45), which is coincidental with substantial elevation of intracellular Ca2+ levels. These findings reveal a set of cellular changes that may represent the earliest markers of in vitro hESC transition to an epithelial phenotype, before the induction of gene expression networks that guide hESC differentiation. Moreover, we hypothesize that these events may be common during the primary steps of hESC commitment to functionally varied epithelial tissue derivatives of different embryological origins.
Stem cells and development 08/2011; 21(8):1250-63. · 4.15 Impact Factor
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ABSTRACT: Transcriptional noise has an important role in generating diversity in cellular populations that are seemingly identical. As this noise stems from the inherent stochasticity of gene expression, it has been unclear whether it is directly controlled. Dig1, a regulator of the budding yeast mating pathway, is now shown to prevent transcriptional noise by regulating the spatial organization of downstream gene targets.
Nature Cell Biology 10/2010; 12(10):929-31. · 19.49 Impact Factor
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ABSTRACT: The nuclei of differentiating cells exhibit several fundamental principles of self-organization. They are composed of many dynamical units connected physically and functionally to each other--a complex network--and the different parts of the system are mutually adapted and produce a characteristic end state. A unique cell-specific signature emerges over time from complex interactions among constituent elements that delineate coordinate gene expression and chromosome topology. Each element itself consists of many interacting components, all dynamical in nature. Self-organizing systems can be simplified while retaining complex information using approaches that examine the relationship between elements, such as spatial relationships and transcriptional information. These relationships can be represented using well-defined networks. We hypothesize that during the process of differentiation, networks within the cell nucleus rewire according to simple rules, from which a higher level of order emerges. Studying the interaction within and among networks provides a useful framework for investigating the complex organization and dynamic function of the nucleus.
Molecular Systems Biology 07/2010; 6:395. · 8.63 Impact Factor
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ABSTRACT: The eukaryotic nucleus is functionally organized. Gene loci, for example, often reveal altered localization patterns according to their developmental regulation. Whole chromosomes also demonstrate non-random nuclear positions, correlated with inherent characteristics such as gene density or size. Given that hundreds to thousands of genes are coordinately regulated in any given cell type, interest has grown in whether chromosomes may be specifically localized according to gene regulation. A synthesis of the evidence for preferential chromosomal organization suggests that, beyond basic characteristics, chromosomes can assume positions functionally related to gene expression. Moreover, analysis of total chromosome organization during cellular differentiation indicates that unique chromosome topologies, albeit probabilistic, in effect define a cell lineage. Future work with new techniques, including the advanced forms of the chromosome conformation capture (3C), and the development of next-generation whole-genome imaging approaches, will help to refine our view of chromosomal organization. We suggest that genomic organization during cellular differentiation should be viewed as a dynamic process, with gene expression patterns leading to chromosome associations that feed back on themselves, leading to the self-organization of the genome according to coordinate gene regulation.
Current opinion in cell biology 06/2010; 22(3):314-9. · 14.15 Impact Factor
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ABSTRACT: Although the importance of chromosome organization during mitosis is clear, it remains to be determined whether the nucleus assumes other functionally relevant chromosomal topologies. We have previously shown that homologous chromosomes have a tendency to associate during hematopoiesis according to their distribution of coregulated genes, suggesting cell-specific nuclear organization. Here, using the mathematical approaches of distance matrices and coupled oscillators, we model the dynamic relationship between gene expression and chromosomal associations during the differentiation of a multipotential hematopoietic progenitor. Our analysis reveals dramatic changes in total genomic order: Commitment of the progenitor results in an initial increase in entropy at both the level of gene coregulation and chromosomal organization, which we suggest represents a phase transition, followed by a progressive decline in entropy during differentiation. The stabilization of a highly ordered state in the differentiated cell types results in lineage-specific chromosomal topologies and is related to the emergence of coherence-or self-organization-between chromosomal associations and coordinate gene regulation. We discuss how these observations may be generally relevant to cell fate decisions encountered by progenitor/stem cells.
Proceedings of the National Academy of Sciences 04/2009; 106(16):6679-84. · 9.68 Impact Factor
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ABSTRACT: Gene loci are found in nuclear subcompartments that are related to their expression status. For instance, silent genes are often localized to heterochromatin and the nuclear periphery, whereas active genes tend to be found in the nuclear center. Evidence also suggests that chromosomes may be specifically positioned within the nucleus; however, the nature of this organization and how it is achieved are not yet fully understood. To examine whether gene regulation is related to a discernible pattern of genomic organization, we analyzed the linear arrangement of co-regulated genes along chromosomes and determined the organization of chromosomes during the differentiation of a hematopoietic progenitor to erythroid and neutrophil cell types. Our analysis reveals that there is a significant tendency for co-regulated genes to be proximal, which is related to the association of homologous chromosomes and the spatial juxtaposition of lineage-specific gene domains. We suggest that proximity in the form of chromosomal gene distribution and homolog association may be the basis for organizing the genome for coordinate gene regulation during cellular differentiation.
PLoS Biology 12/2007; 5(11):e309. · 11.45 Impact Factor
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ABSTRACT: When considering the daunting complexity of eukaryotic genomes, some comfort can be found in the fact that the human genome may contain only 30,000 to 40,000 genes. Moreover, growing evidence suggests that genomes may be organized in such a way as to take advantage of space. A gene's location in the linear DNA sequence and its position in the three-dimensional nucleus can both be important in its regulation. Contrary to prevailing notions in this postgenomic era, the bacteriophage lambda, a paragon of simplicity, may still have a few things to teach us with respect to these facets of nonrandom genomes.
Science 11/2004; 306(5696):644-7. · 31.20 Impact Factor
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ABSTRACT: The extent to which the nucleus is functionally organized has broad biological implications. Evidence supports the idea that basic nuclear functions, such as transcription, are structurally integrated within the nucleus. Moreover, recent studies indicate that the linear arrangement of genes within eukaryotic genomes is nonrandom. We suggest that determining the relationship between nuclear organization and the linear arrangement of genes will lead to a greater understanding of how transcriptomes, dedicated to a particular cellular function or fate, are coordinately regulated. Current network theories may provide a useful framework for modeling the inherent complexity the functional organization of the nucleus.
Genes & Development 07/2004; 18(12):1371-84. · 11.66 Impact Factor
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ABSTRACT: Recent studies of nuclear organization have shown an apparent correlation between the localization of genes within the interphase nucleus and their transcriptional status. In several instances, actively transcribed gene loci have been found significantly looped away from their respective chromosome territories (CTs), presumably as a result of their expression. Here, we show evidence that extrusion of a gene locus from a CT by itself is not necessarily indicative of transcriptional activity, but also can reflect a poised state for activation. We found the murine and a wild-type human beta-globin locus looped away from their CTs at a high frequency only in a proerythroblast cell background, prior to the activation of globin transcription. Conversely, a mutant allele lacking the locus control region (LCR), which is required for high-level globin expression, was mostly coincident with the CT. The LCR may thus be responsible for the localization of the globin locus prior to activation. Replacement of the LCR with a B-cell-specific regulatory element, while also extruding the globin locus, brought it closer to the repressive centromeric heterochromatin compartment. We therefore suggest that the looping of gene loci from their CTs may reflect poised and repressed states, as well as the previously documented transcriptionally active state.
Chromosome Research 02/2003; 11(5):513-25. · 3.09 Impact Factor
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ABSTRACT: The interchromosome domain (ICD) model proposes that genes are selectively positioned at the surfaces of chromosome territories to facilitate their regulation. A paper in the May 13 issue of the Journal of Cell Biology provides evidence that supports a reinterpretation of this model.
Developmental Cell 07/2002; 2(6):690-2. · 14.03 Impact Factor
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ABSTRACT: Immunoglobulin (Ig) loci are selectively activated for transcription and rearrangement during B lymphocyte development. Using fluorescence in situ hybridization, we show that Ig heavy (H) and Igkappa loci are preferentially positioned at the nuclear periphery in hematopoietic progenitors and pro-T cells but are centrally configured in pro-B nuclei. The inactive loci at the periphery do not associate with centromeric heterochromatin. Upon localization away from the nuclear periphery in pro-B cells, the IgH locus appears to undergo large-scale compaction. We suggest that subnuclear positioning represents a novel means of regulating transcription and recombination of IgH and Igkappa loci during lymphocyte development.
Science 05/2002; 296(5565):158-62. · 31.20 Impact Factor
