Gene expression profile at single cell level of NKX6.1- and -NEUROG3 expressing cells and their progeny derived from human induced pluripotent stem cells
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE202092 | GEO | 2023/08/09
REPOSITORIES: GEO
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Project description:Despite this critical role in islet cell development, the precise function and downstream genetic programs regulated directedly by NEUROG3 remain elusive. We therefore mapped genome-wide NEUROG3 occupancy in human induced pluripotent stem cell (iPSC)-derived endocrine progenitors and determined NEUROG3 dependency of associated genes to uncover direct targets. To this aim, we generated a novel hiPSC line (NEUROG3-HA-P2A-Venus), where NEUROG3 is HA-tagged and fused to a self-cleaving fluorescent VENUS reporter. We used the CUT&RUN technique, an alternative method to CHIP-seq allowing transcription factor profiling from a low cell number, to map NEUROG3 occupancy and epigenetic marks in pancreatic endocrine progenitors (PEP) differentiated from this hiPSC line. To optimize the stringency and relevance of NEUROG3 binding sites, we focused our analysis on regions identified both with HA and NEUROG3 antibodies and integrated NEUROG3 occupancy data with chromatin status and gene expression in PEPs and their NEUROG3-dependence. Mapping of NEUROG3 genome occupancy in PEPs uncovers an unexpectedly broad, direct control of the endocrine gene regulatory network (GRN) and raises novel hypotheses on how this master regulator controls islet and beta cell differentiation.
2021-08-04 | GSE171963 | GEO
Project description:Despite this critical role in islet cell development, the precise function and downstream genetic programs regulated directedly by NEUROG3 remain elusive. We therefore mapped genome-wide NEUROG3 occupancy in human induced pluripotent stem cell (iPSC)-derived endocrine progenitors and determined NEUROG3 dependency of associated genes to uncover direct targets. To this aim, we generated a novel hiPSC line (NEUROG3-HA-P2A-Venus), where NEUROG3 is HA-tagged and fused to a self-cleaving fluorescent VENUS reporter. We used the CUT&RUN technique, an alternative method to CHIP-seq allowing transcription factor profiling from a low cell number, to map NEUROG3 occupancy and epigenetic marks in pancreatic endocrine progenitors (PEP) differentiated from this hiPSC line. To optimize the stringency and relevance of NEUROG3 binding sites, we focused our analysis on regions identified both with HA and NEUROG3 antibodies and integrated NEUROG3 occupancy data with chromatin status and gene expression in PEPs and their NEUROG3-dependence. Mapping of NEUROG3 genome occupancy in PEPs uncovers an unexpectedly broad, direct control of the endocrine gene regulatory network (GRN) and raises novel hypotheses on how this master regulator controls islet and beta cell differentiation.
2021-08-04 | GSE179261 | GEO
Project description:Analysis of FACS-sorted intestinal Neurog3+/+ and Neurog3+/- cells from Neurog3 fluorencent reporter mice carrying two-(+/+) or one- (+/-) Neurog3 alleles. In the intestine, Neurog3 is an enteroendocrine (EE) lineage determinating transcription factor that transiently expressed in early EEC progenitors. By comparison the molecular profiles of Neurog3+/+ and Neurog3+/- cells, we hypothesized that Neurog3 gene dosage regulates the allocation of EE progenitor fates towards EEC vs. goblet cells.
2020-08-13 | GSE149203 | GEO
Project description:Atoh8 is a transcription factor of the basic-helix-loop-helix (bHLH) family that is expressed in multiple tissues during embryonic development but whose specific functions remain unknown. The gene encoding Atoh8 is induced by various lineage- determining bHLH transcription factors in cell culture, suggesting a possible common role of this gene in multiple bHLH-driven differentiation programs. In the pancreas, the pro-endocrine bHLH factors Neurogenin3 (Neurog3) and NeuroD1 activate Atoh8 expression. Moreover, Atoh8 is expressed in the embryonic pancreas confirming its participation in the pancreatic transcriptional cascade. This work aims at gaining insight into the molecular function of Atoh8 during the endocrine differentiation program initiated by Neurog3 in the pancreas. To this aim, we have generated a recombinant adenovirus encoding an Atoh8-specific shRNA (Ad-shAtoh8) and used it to down-regulate expression of the Atoh8 gene in Neurog3-expressing pancreatic ductal cells (mPAC), a cellular model of endocrine cell differentiation. Thus, we have compared global changes in gene expression profiles between cells treated with Ad-Neurog3+shControl and cells treated with Ad-Neurog3+shAtoh8 using Affymetrix microarrays. Our results show that Atoh8 silencing significantly affects the expression of 293 genes in Neurog3-expressing mPAC cells. Gene Ontology analysis has revealed cell cycle as the biological function most significantly represented among the modified genes. These results uncover a potential function of Math6 as a regulator of cell cycle progression and provide novel insights into the link between Neurog3 and the regulation of the cell cycle.
2019-01-05 | GSE43537 | GEO
Project description:Nkx6.1 target genes were identified in mature pancreatic islets by comparing gene expression in conditional Nkx6.1-ablated islets versus control islets using microarray analysis.
2013-09-12 | GSE40470 | GEO
Project description:Nkx6.1 target genes were identified in mature pancreatic islets by comparing gene expression in conditional Nkx6.1-ablated islets versus control islets using microarray analysis. Nkx6.1 was conditionally ablated in mature pancreatic islets by recombination of a Nkx6.1-flox allele using the tamoxifen-inducible Pdx1-CreERTM allele (Gu et al 2002). Mice were injected with 2 mg/25 g tamoxifen in corn oil four times between 4 and 6 weeks of age. Islets were isolated after the final tamoxifen injection. Total RNA was isolated and pooled from pancreata of 6 week old Nkx6.1fl/-;Pdx1-CreERTM (mutant) versus Nkx6.1fl/+;Pdx1-CreERTM (control) littermates for 3 biological replicates.
2013-09-12 | E-GEOD-40470 | biostudies-arrayexpress
Project description:To discover cells with mixed enteroendocrine/Paneth/goblet features arising from Neurog3+ progenitors in WT and NFKO mice, we used single-cell RNA sequencing (scRNA-seq) to analyze the diversity of Neurog3 derived cells in the small intestine.
2022-11-06 | GSE201776 | GEO
Project description:Atoh8 is a transcription factor of the basic-helix-loop-helix (bHLH) family that is expressed in multiple tissues during embryonic development but whose specific functions remain unknown. The gene encoding Atoh8 is induced by various lineage- determining bHLH transcription factors in cell culture, suggesting a possible common role of this gene in multiple bHLH-driven differentiation programs. In the pancreas, the pro-endocrine bHLH factors Neurogenin3 (Neurog3) and NeuroD1 activate Atoh8 expression. Moreover, Atoh8 is expressed in the embryonic pancreas confirming its participation in the pancreatic transcriptional cascade. This work aims at gaining insight into the molecular function of Atoh8 during the endocrine differentiation program initiated by Neurog3 in the pancreas. To this aim, we have generated a recombinant adenovirus encoding an Atoh8-specific shRNA (Ad-shAtoh8) and used it to down-regulate expression of the Atoh8 gene in Neurog3-expressing pancreatic ductal cells (mPAC), a cellular model of endocrine cell differentiation. Thus, we have compared global changes in gene expression profiles between cells treated with Ad-Neurog3+shControl and cells treated with Ad-Neurog3+shAtoh8 using Affymetrix microarrays. Our results show that Atoh8 silencing significantly affects the expression of 293 genes in Neurog3-expressing mPAC cells. Gene Ontology analysis has revealed cell cycle as the biological function most significantly represented among the modified genes. These results uncover a potential function of Math6 as a regulator of cell cycle progression and provide novel insights into the link between Neurog3 and the regulation of the cell cycle. mPAC cells (mouse pancreatic duct cells) overexpressing Neurogenin3 were divided in 2 groups according to the shRNA encoded by the recombinant adenoviruses used: 1) Scramble shRNA (control for the infection effect), 2) Atonal Homolog 8-specific shRNA. Experiment was performed three independent times (3 independent biological replicates).
2014-10-10 | E-GEOD-43536 | biostudies-arrayexpress
Project description:Loss of functional β-cell mass is a hallmark of Type 1 and Type 2 diabetes, and methods for restoring these cells are needed. We have previously reported that overexpression of the homeodomain transcription factor Nkx6.1 in rat pancreatic islets induces β-cell proliferation and enhances glucose-stimulated insulin secretion, but the pathway by which Nkx6.1 activates β-cell expansion has not been defined. Here we demonstrate that Nkx6.1 induces expression of the Nr4a1 and Nr4a3 orphan nuclear receptors, and that these factors are both necessary and sufficient for Nkx6.1-mediated β-cell proliferation. Consistent with this finding, global knockout of Nr4a1 results in a decrease in β-cell area in neonatal and young mice. Overexpression of Nkx6.1 and the Nr4a receptors results in increased expression of key cell cycle inducers E2F1 and cyclin E1. Furthermore, Nkx6.1 and Nr4a receptors induce components of the anaphase-promoting complex, including Ube2c, resulting in degradation of the cell cycle inhibitor p21CIP1. These studies identify a new bipartite pathway for activation of β-cell proliferation, suggesting several new targets for expansion of functional β-cell mass.
2014-02-18 | GSE55077 | GEO
Project description:Loss of functional beta-cell mass is a hallmark of Type 1 and Type 2 diabetes, and methods for restoring these cells are needed. We have previously reported that overexpression of the homeodomain transcription factor Nkx6.1 in rat pancreatic islets induces beta-cell proliferation and enhances glucose-stimulated insulin secretion, but the pathway by which Nkx6.1 activates beta-cell expansion has not been defined. Here we demonstrate that Nkx6.1 induces expression of the Nr4a1 and Nr4a3 orphan nuclear receptors, and that these factors are both necessary and sufficient for Nkx6.1-mediated beta-cell proliferation. Consistent with this finding, global knockout of Nr4a1 results in a decrease in beta-cell area in neonatal and young mice. Overexpression of Nkx6.1 and the Nr4a receptors results in increased expression of key cell cycle inducers E2F1 and cyclin E1. Furthermore, Nkx6.1 and Nr4a receptors induce components of the anaphase-promoting complex, including Ube2c, resulting in degradation of the cell cycle inhibitor p21CIP1. These studies identify a new bipartite pathway for activation of beta-cell proliferation, suggesting several new targets for expansion of functional beta-cell mass. We set up a microarray using primary rat islets that were left untreated or transduced with adenoviruses overexpressing betagal or Nkx6.1 for 48 h.
2014-02-18 | E-GEOD-55077 | biostudies-arrayexpress