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The transcription factor Isl1 is associated with multiple signalling pathways important during genital development

ABSTRACT: To identify regulatory targets of the transcription factor Isl1 in the developing mouse genitalia, we performed ChIP-Seq using an anti-ISL1 rabbit monoclonal antibody on pooled chromatin isolated from multiple E14.5 mouse embryonic genital tubercles (GT). Our analysis identified more than 7,054 genomic regions enriched in both of two replicates. The majority of these binding sites are located more than 5 kb from the closest annotated transcriptional start site (TSS), with nearly one third of the binding sites more than 50kb away from the nearest TSS. A de novo motif search using HOMER demonstrated that GT Isl1 ChIP-seq peaks are enriched for a motif that matches the consensus sequence bound by Isl1 in other tissues. When compared to active enhancers in the GT identified using H3K27ac ChIP-Seq, we found that Isl1 binding sites are strongly enriched in the GT compared to other embryonic tissues. Further analysis of the location of Isl1 peaks revealed a strong association with genes implicated in limb and urogenital development. We conclude that there is a strong association between Isl1 regulatory activity and genes involved in genital development.

ORGANISM(S): Mus musculus

PROVIDER: GSE91082 | GEO | 2017/11/08

SECONDARY ACCESSION(S): PRJNA356836

REPOSITORIES: GEO

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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. 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).

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The transcription factor Isl1 is associated with multiple signalling pathways important during genital development

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