Mus musculus
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PROVIDER: PRJNA219490 | ENA |
REPOSITORIES: ENA
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Project description:The motor neuron (MN)–hexamer complex consisting of LIM homeobox 3, Islet-1, and nuclear LIM interactor is a key determinant of motor neuron specification and differentiation. To gain insights into the transcriptional network in motor neuron development, we performed a genome-wide ChIP-sequencing analysis and found that the MN–hexamer directly regulates a wide array of motor neuron genes by binding to the HxRE (hexamer response element) shared among the target genes. Interestingly, STAT3-binding motif is highly enriched in the MN–hexamer–bound peaks in addition to the HxRE. We also found that a transcriptionally active form of STAT3 is expressed in embryonic motor neurons and that STAT3 associates with the MN–hexamer, enhancing the transcriptional activity of the MN–hexamer in an upstream signal-dependent manner. Correspondingly, STAT3 was needed for motor neuron differentiation in the developing spinal cord. Together, our studies uncover crucial gene regulatory mechanisms that couple MN–hexamer and STAT-activating extracellular signals to promote motor neuron differentiation in vertebrate spinal cord.
2013-09-19 | GSE50993 | GEO
Project description:The motor neuron (MN)–hexamer complex consisting of LIM homeobox 3, Islet-1, and nuclear LIM interactor is a key determinant of motor neuron specification and differentiation. To gain insights into the transcriptional network in motor neuron development, we performed a genome-wide ChIP-sequencing analysis and found that the MN–hexamer directly regulates a wide array of motor neuron genes by binding to the HxRE (hexamer response element) shared among the target genes. Interestingly, STAT3-binding motif is highly enriched in the MN–hexamer–bound peaks in addition to the HxRE. We also found that a transcriptionally active form of STAT3 is expressed in embryonic motor neurons and that STAT3 associates with the MN–hexamer, enhancing the transcriptional activity of the MN–hexamer in an upstream signal-dependent manner. Correspondingly, STAT3 was needed for motor neuron differentiation in the developing spinal cord. Together, our studies uncover crucial gene regulatory mechanisms that couple MN–hexamer and STAT-activating extracellular signals to promote motor neuron differentiation in vertebrate spinal cord. To explain our experimental scheme briefly, we are interested in finding target sites for the dimer of transcription factors Isl1 and Lhx3. To mimic the biological activity of Isl1/Lhx3 dimer, we made Isl1-Lhx3 fusion and found that Isl1-Lhx3 has a potent biological activity in multiple systems (i.e. generation of ectopic motor neurons). Then we made ES cell line that induces Flag-tagged Isl1-Lhx3 expression upon Dox treatment. These *mouse* ES cells differentiate to motor neurons (iMN-ESCs) when treated with Dox following EB formation. To identify genomic binding sites of Isl1-Lhx3 (Flag-tagged), we performed ChIP with Flag antibody (pull down of Flag-Isl1-Lhx3) in ES cells treated with Dox. ChIP with Flag antibody in ES cells treated with vehicle (no Dox) was done as a negative control in parallel, and sequenced along with +Dox sample. We have done these experiments twice (two sets).
2013-09-19 | E-GEOD-50993 | biostudies-arrayexpress
Project description:Temporal control of species-specific motor neuron differentiation
| PRJNA590754 | ENA
Project description:Combinatorial transcription codes generate the myriad of cell types during development, and thus likely provide crucial insights into directed differentiation of stem cells to a specific cell type. The LIM-complex composed of Isl1 and Lhx3 directs the specification of spinal motor neurons (MNs) in embryos. Here, we report that Isl1-Lhx3, a LIMcomplex-mimicking fusion, induces a signature of MN transcriptome and concomitantly suppresses interneuron differentiation programs, thereby serving as a potent and specific inducer of MNs in stem cells. We show that an equimolar ratio of Isl1 and Lhx3 and the LIM-domain of Lhx3 are crucial for generating MNs without upregulating interneuron genes. These led us to design Isl1-Lhx3, which maintains the desirable 1:1 ratio of Isl1 and Lhx3 and the LIM-domain of Lhx3. Isl1-Lhx3 drives MN differentiation with high specificity and efficiency in the spinal cord and embryonic stem cells, bypassing the need for sonic hedgehog. RNA-seq analysis revealed that Isl1-Lhx3 induces the expression of a battery of MN genes that control various functional aspects of MNs, while suppressing key interneuron genes. Our studies uncover a highly efficient method for directed MN generation and MN gene networks. Our results also demonstrate a general strategy of utilizing embryonic transcription complexes for producing specific cell types from stem cells. Examine the RNA expression profiles of inducible motor neurons-embryonic stem cells (iMN-ESCs) with or without Dox treatment, with biological duplicates.
2012-02-03 | E-GEOD-35510 | biostudies-arrayexpress
Project description:Temporal control of species-specific motor neuron differentiation [Mm]
| PRJNA590756 | ENA
Project description:Temporal control of species-specific motor neuron differentiation [Hs]
| PRJNA590757 | ENA
Project description:The fidelity of motor control requires the precise positional arrangement of motor pools and the establishment of synaptic connections between these pools. In the developing spinal cord, motor nerves project to specific target muscles and receive proprioceptive input from the muscles via the sensorimotor circuit. LIM-homeodomain transcription factors are known to successively restrict specific motor neuronal fates during neural development; however, it remains unclear to what extent they contribute to limb-based motor pools and locomotor circuits. Here, we showed in mice that deletion of Isl2 resulted in scattered motor pools, primarily in the median motor column and lateral LMC (LMCl) populations, and lacked Pea3 expression in the hindlimb motor pools, accompanied by reduced terminal axon branching and disorganized neuromuscular junctions. Transcriptomic analysis of Isl2-deficient spinal cords revealed that a variety of genes involved in motor neuron differentiation, axon development, and synapse organization were downregulated in hindlimb motor pools. Moreover, the loss of Isl2 impaired sensorimotor connectivity and hindlimb locomotion. Together, our studies indicate that Isl2 plays a critical role in organizing motor pool position and sensorimotor circuits in hindlimb motor pools.
2023-11-01 | GSE217297 | GEO
Project description:Generic spinal motor neuron identity is specified by cooperative binding of selector transcription factors to motor neuron specific enhancers. Whether these enhancers remain active to maintain the motor neuron expression program following downregulation of selector factors in maturing motor neurons remains unknown. We demonstrate that enhancers established by selector genes are highly transient during motor neuron differentiation. The chromatin immunoprecipitation-exonuclease (ChIP-exo) assay reveals that Isl1 is anchored to nascent motor neuron enhancers through protein-protein interaction. Stage-specific recruitment of transcription factors correlates with active enhancer marks. The majority of genes contain distinct early and late enhancers suggesting that the motor neuron expression program is maintained by a dynamic regulatory landscape.
2016-12-05 | GSE79561 | GEO
Project description:During development, two cell-types born from closely related progenitor pools often express the identical transcriptional regulators despite their completely distinct characteristics. This phenomenon highlights the necessity of the mechanism that operates to segregate the identities of the two cell-types throughout differentiation after initial fate commitment. To understand this mechanism, we investigated the fate specification of spinal V2a interneurons, which share important developmental genes with motor neurons (MNs). Here we demonstrate that the paired homeodomain factor Chx10 functions as a critical determinant for V2a fate and is required to consolidate V2a identity in postmitotic neurons. Chx10 actively promotes V2a fate, downstream of the LIM-homeodomain factor Lhx3, while concomitantly suppressing MN developmental program by preventing the MN-specific transcription complex from binding and activating MN genes. This dual activity enables Chx10 to effectively separate V2a and MN pathways. Together, our study uncovers a widely applicable gene regulatory principle for segregating related cell fates. RNA samples from Chx10-ESC-derived MNs were prepared for sequencing according to the Illumina protocol, and sequenced on the Illumina HiSeq 2000. We will then compare the transcriptome changes between -Dox (no Chx10) and +Dox (Chx10) in order to identify genes rregulated by Chx10.
2016-07-28 | E-GEOD-83873 | biostudies-arrayexpress
Project description:Human pluripotent stem cells are a promising source of diverse cells for developmental studies, cell transplantation, disease modeling, and drug testing. However, their widespread use even for intensely studied cell types like spinal motor neurons, is hindered by the long duration and low yields of existing protocols for in vitro differentiation and by the molecular heterogeneity of the populations generated. We report a combination of small molecules that induce up to 50% motor neurons within 3 weeks from human pluripotent stem cells with defined subtype identities that are relevant to neurodegenerative diseases. Despite their accelerated differentiation, motor neurons expressed combinations of HB9, ISL1 and column-specific markers that mirror those observed in vivo in human fetal spinal cord. They also exhibited spontaneous and induced activity, and projected axons towards muscles when grafted into developing chick spinal cord. Strikingly, this novel protocol preferentially generates motor neurons expressing markers of limb-innervating lateral motor column motor neurons (FOXP1+/LHX3-). Access to high-yield cultures of human limb-innervating motor neuron subtypes will facilitate in-depth study of motor neuron subtype-specific properties, disease modeling, and development of large-scale cell-based screening assays. We analyze 3 samples including 2 positive samples and 1 negative sample. Descriptions are as follow: a) Positive Sample 1: SHH-derived, day 21 GFP-high FACS purified motor neurons.b) Positive Sample 2: S+P-derived, day 21 GFP-high FACS purified motor neurons. c) Negative: S+P condition, day 21 no GFP FACS purified motor neurons
2013-02-06 | E-GEOD-41795 | biostudies-arrayexpress