Project description:The genomic regulatory programs that underlie human organogenesis are poorly understood. Human pancreas development, in particular, has pivotal implications for pancreatic regeneration, cancer, and diabetes. We have now created maps of transcripts, active enhancers, and transcription factor networks in pancreatic multipotent progenitors obtained from human embryos, or derived in vitro from human embryonic stem cells. This revealed that artificial progenitors recapitulate salient transcriptional and epigenomic features of their natural counterparts. Using this resource, we show that TEAD1, a transcription factor controlled by Hippo signaling, is a core component of the combinatorial code of pancreatic progenitor enhancers. TEAD thus activates genes encoding regulators of signaling pathways and stage-specific transcription factors that are essential for normal pancreas development. Accordingly, chemical and genetic perturbations of TEAD and its coactivator YAP inhibited expression of known regulators such as FGFR2 and SOX9, and suppressed the proliferation and expansion of mouse and zebrafish pancreatic progenitors. These findings provide a resource of active enhancers and transcripts in human pancreatic multipotent progenitors, and uncover a central role of TEAD and YAP as signal-responsive regulators of the transcriptional program of early pancreas development.
Project description:Our lab identified Sox9 as a specific marker and maintenance factor of mouse pancreatic progenitors (Seymour et al., PNAS, 2007). Here was wanted to identify direct targets of Sox9 in pancreatic progenitors. However, due to the limited number of pancreatic progenitors in the developing mouse, we used in vitro derived pancreatic progenitors to determine direct targets of Sox9. We performed ChIP-seq analysis for Sox9 and determined its direct targets in the human genome. Using pancreatic progenitors that were derived from human embryonic stem cells, we were able to successfully find targets that were pancreas specific as well as those targets that are important in other endodermal lineages. Overall design: We differentiated CyT49 human embryonic stem cells into pancreatic progenitors as previously described (Kroon et al., 2008; Schulz et al., 2012). Using a Sox9 antibody, we performed ChIP-seq in the pancreatic progenitors. We sequenced the samples using the Illumina platform and mapped the reads to the mm9 version of the mouse genome. Using HOMER analysis software and the UCSC genome browser, we found direct targets of Sox9 in human embryonic stem cell derived pancreatic progenitors.
Project description:Active regulatory regions in the human embryonic pancreatic progenitors were profiled by integration of transcription factor and histone modification ChIP-seq datasets. These were obtained from pancreatic progenitor cells derived in vitro from human embryonic stem cells. The purpose of this work was to study the epigenomic mechanisms involved in pancreas development.
Project description:Chavez2009 - a core regulatory network of OCT4 in human embryonic stem cells
A core OCT4-regulated network has been identified as a test case, to analyase stem cell characteristics and cellular differentiation.
This model is described in the article:
In silico identification of a core regulatory network of OCT4 in human embryonic stem cells using an integrated approach.
Chavez L, Bais AS, Vingron M, Lehrach H, Adjaye J, Herwig R
BMC Genomics, 2009, 10:314
BACKGROUND: The transcription factor OCT4 is highly expressed in pluripotent embryonic stem cells which are derived from the inner cell mass of mammalian blastocysts. Pluripotency and self renewal are controlled by a transcription regulatory network governed by the transcription factors OCT4, SOX2 and NANOG. Recent studies on reprogramming somatic cells to induced pluripotent stem cells highlight OCT4 as a key regulator of pluripotency.
RESULTS: We have carried out an integrated analysis of high-throughput data (ChIP-on-chip and RNAi experiments along with promoter sequence analysis of putative target genes) and identified a core OCT4 regulatory network in human embryonic stem cells consisting of 33 target genes. Enrichment analysis with these target genes revealed that this integrative analysis increases the functional information content by factors of 1.3 - 4.7 compared to the individual studies. In order to identify potential regulatory co-factors of OCT4, we performed a de novo motif analysis. In addition to known validated OCT4 motifs we obtained binding sites similar to motifs recognized by further regulators of pluripotency and development; e.g. the heterodimer of the transcription factors C-MYC and MAX, a prerequisite for C-MYC transcriptional activity that leads to cell growth and proliferation.
CONCLUSION: Our analysis shows how heterogeneous functional information can be integrated in order to reconstruct gene regulatory networks. As a test case we identified a core OCT4-regulated network that is important for the analysis of stem cell characteristics and cellular differentiation. Functional information is largely enriched using different experimental results. The de novo motif discovery identified well-known regulators closely connected to the OCT4 network as well as potential new regulators of pluripotency and differentiation. These results provide the basis for further targeted functional studies.
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Project description:Human pluripotent stem cells were differentiated into PDX1+/NKX6-1+ Pancreatic Progenitors (PPd15 cells), which were subsequently captured and expanded in culture. These culture Pancreatic Progenitors (cPP cells) were capable of self-renewal and could be passaged up to 20 times. Furthermore, cPP cells were capable of differentiation into multiple pancreatic lineages, including c-peptide+ beta-like cells, both in vitro and in vivo.
Project description:We used epigenetic profiling to map active enhancers in the developing human limb and cortex as described in two published studies: Cotney J, Leng J, Yin J, Reilly SK et al. The evolution of lineage-specific regulatory activities in the human embryonic limb. Cell 2013;154(1):185-96. Reilly SK, Yin J, Ayoub AE, Emera D et al. Evolutionary changes in promoter and enhancer activity during human corticogenesis. Science 2015;347(6226):1155-9. We also used ChIP-seq to map binding sites for the chromatin modifier CHD8 in the developing human brain, as described in one published study: Cotney J, Muhle RA, Sanders SJ, Liu L et al. The autism-associated chromatin modifier CHD8 regulates other autism risk genes during human neurodevelopment. Nat Commun 2015;6:6404. We are also depositing primary human sequence reads related to processed datasets in the Gene Expression Omnibus under the following accession numbers: GSE42413(Cotney et al. 2013); GSE63649(Reilly... (for more see dbGaP study page.)
Project description:To guide the beta cell differentiation process in vitro, a complete understanding of the transcriptome and their regulatory network during the differentiation process is essential. Using RNA-seq, we have performed the transcriptome profiling of human embryonic stem cells (ESCs), purified ESC-derivate definitive endoderm (DE), pancreatic progenitors (PP), as well as sorted human primary pancreatic alpha cells, beta cells and exocrine cells.