Controlled induction of 49 transcription factors in transgenic mouse ES cell lines
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE72350 | GEO | 2016/07/21
SECONDARY ACCESSION(S): PRJNA293862
REPOSITORIES: GEO
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Project description:Controlled induction of individual transcription factors (TFs) in embryonic stem cells (ESCs) with subsequent global gene expression profiling is a promising approach for identification of downstream genes regulated by these TFs in a relatively promiscuous epigenetic background. Here we generated and characterized mouse ESCs clones with 49 doxycycline-controllable TFs as a continuation of the NIA Mouse ESC Bank project. Together with the previous clone libraries, the project now comprises 186 TFs (ca. 10% of all TFs in the genome). Induction of individual TFs shifts the transcriptome towards specific differentiation fates (e.g., neural for Myt1, St18; fibroblasts and osteoblasts for Pitx1, Pitx2, Barhl2, and Lmx1a; thymocytes for Myb; lymph nodes and spleen for Etv2 and Plac1, and ovary for Pitx1, Pitx2, and Dmrtc2). Gene set enrichment analysis for expression profile change after induction of TFs provides additional information on the functions and phenotypes associated with these TFs. These data could facilitate methods for generating various types of cells in regenerative medicine as well as ameliorating pathogenic phenotypes in gene therapy. Dox- overexpressing transcription factor vs. Dox+ (control).
2016-07-21 | E-GEOD-72350 | biostudies-arrayexpress
Project description:Islet β-cells from newborn mammals need a maturation process to become mature functional beta cells. The detailed molecular mechanisms were not completely understood. This study was designed to reveal the dynamic gene expression changes during pancreatic beta-cell maturation in postnatal mice. We also want to understand how genetic mutations that impair beta-cell function change the genetic networks involved in the beta-cell maturation process. For these aims, pancreatic beta cells were isolated at P1 islets based on the expression of a MipeGFP transgene in a genetic background with pancreatic specific inactivation of Myt1, Myt1L, and St18 (denoted as MytDelpanc; MipeGFP).
2018-04-01 | E-MTAB-6615 | biostudies-arrayexpress
Project description:Recent developments in molecular programming of mesodiencephalic dopaminergic (mdDA) neurons have led to the identification of many transcription factors playing a role in mdDA specification. LIM homeodomain transcription factor Lmx1a is essential for chick mdDA development, and for the efficient differentiation of ES-cells towards a dopaminergic phenotype. In this study, we aimed towards a more detailed understanding of the subtle phenotype in Lmx1a-dr/dr mice. Therefore, microarray analysis was performed, to elucidate the exact molecular programming underlying the neuronal deficits after loss of Lmx1a. Subsequent expression analysis confirmed that Nurr1 is regulated by Lmx1a, and additional downstream targets were identified, like Pou4f1, Pbx1, Pitx2, C130021l20Rik, Calb2 and Rspo2. In line with a specific, rostral-lateral loss of expression of most of these genes during development, Nurr1 and C130021l20Rik were affected in the SNc of the mature mdDA system. Interestingly, this deficit was marked by the complete loss of the Wnt/b-catenin signaling activator Rspo2 in this domain. Expression analysis in Rspo2-/- embryos revealed affected mdDA neurons, partially phenocopying the Lmx1a mutant. Together, in this study we reveal that Lmx1a is essential for a rostral-lateral subset of the mdDA neuronal field, where it might serve a critical function in modulating proliferation and differentiation of mdDA progenitors through the activation of the Wnt activator Rspo2. Microarray expression study comparing 4 samples of homozygous LMX1A dr/dr mice, midbrain E12.5 with a pooled sample of their wt/wt littermates. Two samples were analyzed in opposite dye orientation.
2013-12-13 | E-GEOD-45831 | biostudies-arrayexpress
Project description:Recent developments in molecular programming of mesodiencephalic dopaminergic (mdDA) neurons have led to the identification of many transcription factors playing a role in mdDA specification. LIM homeodomain transcription factor Lmx1a is essential for chick mdDA development, and for the efficient differentiation of ES-cells towards a dopaminergic phenotype. In this study, we aimed towards a more detailed understanding of the subtle phenotype in Lmx1a-dr/dr mice. Therefore, microarray analysis was performed, to elucidate the exact molecular programming underlying the neuronal deficits after loss of Lmx1a. Subsequent expression analysis confirmed that Nurr1 is regulated by Lmx1a, and additional downstream targets were identified, like Pou4f1, Pbx1, Pitx2, C130021l20Rik, Calb2 and Rspo2. In line with a specific, rostral-lateral loss of expression of most of these genes during development, Nurr1 and C130021l20Rik were affected in the SNc of the mature mdDA system. Interestingly, this deficit was marked by the complete loss of the Wnt/b-catenin signaling activator Rspo2 in this domain. Expression analysis in Rspo2-/- embryos revealed affected mdDA neurons, partially phenocopying the Lmx1a mutant. Together, in this study we reveal that Lmx1a is essential for a rostral-lateral subset of the mdDA neuronal field, where it might serve a critical function in modulating proliferation and differentiation of mdDA progenitors through the activation of the Wnt activator Rspo2.
2013-12-13 | GSE45831 | GEO
Project description:Specific neuronal types derived from embryonic stem cells (ESCs) can facilitate mechanistic studies and potentially aid in regenerative medicine. Existing induction methods, however, mostly rely on the effects of growth factors, which generally tend to result in mixed populations of neurons. Here we report that over-expression of specific transcription factors (TFs) in ESCs can rather guide the differentiation of ESCs towards specific neuron types. Analysis of published data on gene expression changes early (two days) after induction of each of 185 induced TFs implicated candidate TFs for further ESC differentiation studies. After induction for 6 days four of them (Ascl1, Smad7, Nr2f1, and Ascl2) generated a high proportion (>35%) of cells with neural progenitor marker PSA-NCAM and clear neural morphology on day 14. The capacity of these TFs to induce neural differentiation is inferred to be most likely linked to early activation of the Notch signaling pathway. Among the neuron-like cells, GABA-positive cells were most abundant (32-97% for 4 top TFs), whereas Isl1-positive cells and TH-positive cells were less abundant (<12% and <5%, respectively). Enrichment of cells obtained with the induction of Ascl1, Smad7, and Nr2f1 using beads with anti-PSA-NCAM antibody resulted in essentially pure population of neuron-like cells with expression profiles similar to neural tissues and highly expressed markers of GABAergic neurons. A time-course experiment with induction of Ascl1 showed early upregulation of most neural-specific and GABAergic-specific mRNA and miRNAs. We identified mRNA and miRNAs, whose expression depended on the induction of Ascl1, and showed that they were enriched in Ascl1 target genes. In summary, this study indicates that induction of transcription factors is a promising approach to generate candidate specific neural cell types from ESCs. Individual transcription factors (TFs) (Ascl1, Smad7, and Nr2f1) were induced in mouse ESCs to facilitate neural differentiation. Expression of transgenic TFs was repressed by doxycycline (Dox); thus, TFs were induced in Dox- conditions, whereas Dox+ conditions represent control cells with no expression of a transgene. For neural differentiation, cells were cultured 3 days in alpha-MEM medium and then in NeuroCult neural differentiation medium. Cells marked as PSANCAM+ were enriched by magnetic microbeads with anty-PSA-NCAM antibody (Miltenyi Biotec) on day 6 of culturing, whereas remaining cells are marked as PSANCAM-. Both PSANCAM+ and PSANCAM- cells were then cultured for another 8 days (total 14 days in differentiation). A time course experiment with Ascl1 induction did not include cell enrichment procedure. RNA was extracted with either Trizol (invitrogen) or mirVana kit (Thermo Fisher Scientific).
2016-07-21 | E-GEOD-78949 | biostudies-arrayexpress
Project description:Specific neuronal types derived from embryonic stem cells (ESCs) can facilitate mechanistic studies and potentially aid in regenerative medicine. Existing induction methods, however, mostly rely on the effects of growth factors, which generally tend to result in mixed populations of neurons. Here we report that over-expression of specific transcription factors (TFs) in ESCs can rather guide the differentiation of ESCs towards specific neuron types. Analysis of published data on gene expression changes early (two days) after induction of each of 185 induced TFs implicated candidate TFs for further ESC differentiation studies. After induction for 6 days four of them (Ascl1, Smad7, Nr2f1, and Ascl2) generated a high proportion (>35%) of cells with neural progenitor marker PSA-NCAM and clear neural morphology on day 14. The capacity of these TFs to induce neural differentiation is inferred to be most likely linked to early activation of the Notch signaling pathway. Among the neuron-like cells, GABA-positive cells were most abundant (32-97% for 4 top TFs), whereas Isl1-positive cells and TH-positive cells were less abundant (<12% and <5%, respectively). Enrichment of cells obtained with the induction of Ascl1, Smad7, and Nr2f1 using beads with anti-PSA-NCAM antibody resulted in essentially pure population of neuron-like cells with expression profiles similar to neural tissues and highly expressed markers of GABAergic neurons. A time-course experiment with induction of Ascl1 showed early upregulation of most neural-specific and GABAergic-specific mRNA and miRNAs. We identified mRNA and miRNAs, whose expression depended on the induction of Ascl1, and showed that they were enriched in Ascl1 target genes. In summary, this study indicates that induction of transcription factors is a promising approach to generate candidate specific neural cell types from ESCs. Transcription factor Ascl1 was induced in mouse ESCs to facilitate neural differentiation. Expression of transgenic Ascl1 was repressed by doxycycline (Dox); thus, it were induced in Dox- conditions, whereas Dox+ conditions represent control cells with no expression of Ascl1 transgene. For neural differentiation, cells were cultured 3 days in alpha-MEM medium and then - in NeuroCult neural differentiation medium for 2-11 days (total up to 14 days). RNA was extracted with mirVana kit (Thermo Fisher Scientific).
2016-07-21 | E-GEOD-78950 | biostudies-arrayexpress
Project description:Specific neuronal types derived from embryonic stem cells (ESCs) can facilitate mechanistic studies and potentially aid in regenerative medicine. Existing induction methods, however, mostly rely on the effects of growth factors, which generally tend to result in mixed populations of neurons. Here we report that over-expression of specific transcription factors (TFs) in ESCs can rather guide the differentiation of ESCs towards specific neuron types. Analysis of published data on gene expression changes early (two days) after induction of each of 185 induced TFs implicated candidate TFs for further ESC differentiation studies. After induction for 6 days four of them (Ascl1, Smad7, Nr2f1, and Ascl2) generated a high proportion (>35%) of cells with neural progenitor marker PSA-NCAM and clear neural morphology on day 14. The capacity of these TFs to induce neural differentiation is inferred to be most likely linked to early activation of the Notch signaling pathway. Among the neuron-like cells, GABA-positive cells were most abundant (32-97% for 4 top TFs), whereas Isl1-positive cells and TH-positive cells were less abundant (<12% and <5%, respectively). Enrichment of cells obtained with the induction of Ascl1, Smad7, and Nr2f1 using beads with anti-PSA-NCAM antibody resulted in essentially pure population of neuron-like cells with expression profiles similar to neural tissues and highly expressed markers of GABAergic neurons. A time-course experiment with induction of Ascl1 showed early upregulation of most neural-specific and GABAergic-specific mRNA and miRNAs. We identified mRNA and miRNAs, whose expression depended on the induction of Ascl1, and showed that they were enriched in Ascl1 target genes. In summary, this study indicates that induction of transcription factors is a promising approach to generate candidate specific neural cell types from ESCs.
2016-07-21 | GSE78950 | GEO
Project description:Specific neuronal types derived from embryonic stem cells (ESCs) can facilitate mechanistic studies and potentially aid in regenerative medicine. Existing induction methods, however, mostly rely on the effects of growth factors, which generally tend to result in mixed populations of neurons. Here we report that over-expression of specific transcription factors (TFs) in ESCs can rather guide the differentiation of ESCs towards specific neuron types. Analysis of published data on gene expression changes early (two days) after induction of each of 185 induced TFs implicated candidate TFs for further ESC differentiation studies. After induction for 6 days four of them (Ascl1, Smad7, Nr2f1, and Ascl2) generated a high proportion (>35%) of cells with neural progenitor marker PSA-NCAM and clear neural morphology on day 14. The capacity of these TFs to induce neural differentiation is inferred to be most likely linked to early activation of the Notch signaling pathway. Among the neuron-like cells, GABA-positive cells were most abundant (32-97% for 4 top TFs), whereas Isl1-positive cells and TH-positive cells were less abundant (<12% and <5%, respectively). Enrichment of cells obtained with the induction of Ascl1, Smad7, and Nr2f1 using beads with anti-PSA-NCAM antibody resulted in essentially pure population of neuron-like cells with expression profiles similar to neural tissues and highly expressed markers of GABAergic neurons. A time-course experiment with induction of Ascl1 showed early upregulation of most neural-specific and GABAergic-specific mRNA and miRNAs. We identified mRNA and miRNAs, whose expression depended on the induction of Ascl1, and showed that they were enriched in Ascl1 target genes. In summary, this study indicates that induction of transcription factors is a promising approach to generate candidate specific neural cell types from ESCs.
2016-07-21 | GSE78949 | GEO
Project description:Development of the nervous system depends on signaling centers – specialized cellular populations that produce secreted molecules to regulate neurogenesis in the neighboring neuroepithelium. Some signaling centers also generate key types of neurons. The formation of a signaling center involves its induction, the maintenance of expression of its secreted molecules, and cell differentiation and migration events. How these distinct processes are coordinated during signaling center development remains unknown. Here we show that Lmx1a acts as a master regulator to orchestrate the formation and function of the cortical hem (CH), a critical signaling center that controls hippocampus development. Lmx1a co-regulates CH induction, its Wnt signaling, and the differentiation and migration of CH-derived Cajal-Retzius neurons. Combining RNAseq, genetic, and rescue experiments, we identified major downstream genes that mediate distinct Lmx1a-dependent processes. Our work revealed that signaling centers in the mammalian brain employ master regulatory genes and established a framework for analyzing signaling center development.
2023-09-06 | GSE216165 | GEO
Project description:We applied ChIP-seq to explore the effect of missense mutations in TFs on their genome wide binding profile. Using a retroviral expression system in chicken mesenchymal stem cells, we elucidated the mechanism underlying a novel missense mutation in HOXD13 (Q317K) associated with a complex hand and foot malformation phenotype. The glutamine at position 317 (position 50 of the homeodomain) is conserved in most homeodomains, a notable exception being bicoid-type homeodomains that have K at this position. Our results show that the mutation results in a shift in the binding profile of the mutant towards a bicoid/PITX1 motif. Gene expression analysis and functional assays using in vivo overexpression studies confirm that the mutation results in a partial conversion of HOXD13 into a TF with bicoid/PITX1 properties. A similar shift was not observed with the mutation Q317R, which is associated with brachysyndactyly, suggesting that the bicoid/PITX1-shift observed for Q317K might be related to the severe clinical phenotype. For each wt or mutant transcription factor two independent biological replicates were used for ChIP-seq together with corresponding input DNA as reference samples. For expression analysis one RNA sample for each wt and mutant transcription factor was used and compared to the expression level of cells infected with empty vector by RNA-seq.
2013-08-19 | E-GEOD-44799 | biostudies-arrayexpress