RNA-seq of human nociceptor-like sensory neurons with varying Neurogenin 1 induction lengths
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Illumina MiSeq
ORGANISM(S): Homo sapiens
SUBMITTER: Muad Abd El Hay
PROVIDER: E-MTAB-12098 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Induced pluripotent stem cells were differentiated into neural crest progenitors and infected with a lentiviral construct that contains a doxycyclin-inducible Neurogenin 1(Ngn1) cassette. The samples represent RNA-sequencing from initial neural crest stage (day 0) and doxycyclin treatment (days 0 - 10) followed by differentiation without induction of Ngn1 (days 12 till end).
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Project description:Neural stem cells (NSC) with self-renewal and multipotent properties serve as an ideal cell source for transplantation to treat spinal cord injury, stroke, and neurodegenerative diseases. To efficiently induce neuronal lineage cells from NSC for neuron replacement therapy, we should clarify the intrinsic genetic programs involved in a time and place-specific regulation of human NSC differentiation. Recently, we established an immortalized human NSC clone HB1.F3 to provide an unlimited NSC source applicable to genetic manipulation for cell-based therapy. To investigate a role of neurogenin 1 (Ngn1), a proneural basic helix-loop-helix (bHLH) transcription factor, in human NSC differentiation, we established a clone derived from F3 stably overexpressing Ngn1. Genome-wide gene expression profiling identified 250 upregulated genes and 338 downregulated genes in Ngn1-overexpressing F3 cells (F3-Ngn1) versus wild-type F3 cells (F3-WT). Notably, leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5), a novel stem cell marker, showed a robust increase in F3-Ngn1.
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Project description:A genomic overview of in vivo binding of a transcription factor Neurogenin in the ascidian 64-cell stage embryo. Fertilized eggs were electroporated with a construct encoding GFP-tagged Neurogenin. After fixation, chromatin was sonicated to obtain DNA fragments with an average of size of 1kb. The DNA fragments were enriched by immunoprecipitation with an polyclonal antibody specific for GFP (Invitrogen). The immunoprecipitated and WCE (Whole Cell Extract) DNA were amplified by LM-PCR for the Chip analysis. Two independent experiments were performed to reduce experimental errors.
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Project description:To study the early gene expression events involved in neurogenesis, we expressed the bHLH neural determination factor neurogenin 1 in mouse embryonic stem cells. We used microarray to examine the program of gene expression at 24, 48, 72 hours of transgene expression. Mouse embryonic stem cells were engineered to express neurogenin 1 using a tet-sensitive expression system. RNAs from uninduced and induced cells were isolated at 24, 48, and 72 hours of differentiation and subjected to microarray analysis.
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Project description:To study the early gene expression events involved in neurogenesis, we expressed the bHLH neural determination factor neurogenin 1 in mouse embryonic stem cells. We used microarray to examine the program of gene expression at 24, 48, 72 hours of transgene expression.
2013-01-15 | GSE42883 | GEO
Project description:Transcriptional targets of neurogenin (Ngnr1) were identified by over-expression of an inducible form of neurogenin in Xenopus ectodermal explants. The effects of co-expressing the nucleoprotein geminin on Ngnr1-dependent target gene transactivation were defined. Regulating the transition from lineage-restricted progenitors to terminally differentiated cells is a central aspect of nervous system development. Here, we investigated the role of the nucleoprotein geminin in regulating neurogenesis at a mechanistic level during both Xenopus primary neurogenesis and mammalian neuronal differentiation in vitro. The latter work utilized both neural cells derived from embryonic stem and embryonal carcinoma cells in vitro and neural stem cells from mouse forebrain. In all of these contexts, geminin antagonized the ability of neural bHLH transcription factors to activate transcriptional programs promoting neurogenesis. Furthermore, geminin promoted a bivalent chromatin state, characterized by the presence of both activating and repressive histone modifications, at genes encoding transcription factors that promote neurogenesis. This epigenetic state restrains the expression of genes that regulate commitment of undifferentiated stem and neuronal precursor cells to neuronal lineages. Geminin is highly expressed in undifferentiated neuronal precursor cells but is downregulated prior to differentiation. Therefore, these data support a model whereby geminin promotes the neuronal precursor cell state by modulating both the epigenetic status and expression of genes encoding neurogenesis-promoting factors. Additional developmental signals acting in these cells can then control their transition toward terminal neuronal or glial differentiation during mammalian neurogenesis. A dexamethasone-inducible (GR ligand binding domain fused) form of Xenopus neurogenin-related 1, NgnrGR, was over-expressed in Xenopus embryonic ectodermal explants, in the presence or absence of over-expressed geminin. Induction of Ngnr1 activity was used to define direct targets as previously described (EMBO J. 26(24): 5093-5108). The ability of geminin to suppress Ngnr1-dependent transactivation of its target gene programs was determined.
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Project description:Advances in cellular reprogramming and stem cell differentiation now enable ex vivo studies of human neuronal differentiation. However, it remains challenging to elucidate the underlying regulatory programs because differentiation protocols are laborious and often result in low neuron yields. Here, we overexpressed two murine Neurogenin transcription factors in human induced pluripotent stem cells, and obtained neurons with bipolar morphology in four days at greater than 90% purity. The high purity enabled mRNA and microRNA expression profiling during neurogenesis, thus revealing the genetic programs involved in the transition from stem cell to neuron. These profiles were then analyzed to identify the regulatory networks underlying the differentiation of the neurons. Paired end RNA sequencing of iPS cells (PGP1) at 0, 1, 3, and 4 days post- doxycycline induction of murine NGN1 and NGN2. This was done using an Illumina HiSeq, and reads were aligned to hg19
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Project description:Proper nervous system development is required for an organism’s survival and function. Defects in neurogenesis have been linked to neurodevelopmental disorders such as schizophrenia and autism spectrum disorders. Understanding the gene regulatory networks that orchestrate neural development, specifically cascades of proneural transcription factors, can better elucidate which genes are most essential in governing early neurogenesis. Neurogenins are a family of such factors that are both sufficient and necessary for the development of neural sub-types in mice, primarily through the regulation of other factors, particularly neurod. The objective of this study was to evaluate previously established regulatory targets of neurogenin (ngn-1) and to identify unknown downstream targets, using the nematode Caenorhabditis elegans as a model for these studies. We find that in C. elegans, neurogenin is required for axon outgrowth, cell fate specification, and epithelial integrity during embryonic development. Using RNA sequencing and comparative transcriptome analysis we found that ngn-1 acts primarily to repress transcription, facilitating proper embryogenesis. We also identified specific candidates for activation by ngn-1, including hlh-34(npas1) and unc-42(prop1), which we further validated using genetic methods. Our results identify novel pathways connecting ngn-1 to known terminal regulators, which maintain cell fate of terminally differentiated neural subtypes, and align neurogenins within the larger context of proneural transcription factors.
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