Gallus gallus
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA308306 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR3092120.fastq.gz | Fastqsanger.gz | |||
| SRR3092121.fastq.gz | Fastqsanger.gz | |||
| SRR3092118.fastq.gz | Fastqsanger.gz | |||
| SRR3092119.fastq.gz | Fastqsanger.gz |
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Project description:To explore what FOXL2-dependent changes of gene expression observed in our RNA-seq data were directly related to FOXL2 binding, we performed ChIP-seq analysis in FOXL2-expressing granulosa cells (two data points). To explore the potential co-regulation between FOXL2 and ESR2 of several targets, we knocked-down Esr2 in granulosa cells and performed a ChIP-seq under the same conditions as those previously described for FOXL2.
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Project description:FOXL2 is a transcription factor that plays a key role in sex determination, ovary development and maintenance. Mutations related to this gene have been described in syndromes involving premature ovarian failure and granulosa cell tumors. This kind of rare cancer (less than 5% of diagnosed ovarian cancers) has been causally associated with the FOXL2 c.402C>G, p.C134W mutation in 97% of the adult cases (AGCTs). In this study, we have used CRISPR technology to specifically eliminate the FOXL2 c.402C>G mutation in granulosa tumor cells. Our results indicate that this Cas9-mediated strategy allows the specific elimination of the mutation with no activity on the wild type allele. Granulosa cells depleted on FOXL2 c.402C>G show a reduced malignant phenotype. Specifically, we detect changes in cell proliferation, invasion, and cell death levels. In addition, we show that granulosa tumor cells become more susceptible to Dasatinib and Ketoconazole treatments when FOXL2 c.402C>G allele is eliminated. Our transcriptomic and proteomic analyses indicate that CRISPR-modified granulosa tumor cells significantly change their expression signature towards a wild type like phenotype. Finally, this expression signature has led us to discover new compounds with antiproliferative and proapoptotic effects on granulosa cell tumor cells. Our results demonstrate the potential of CRISPR for specifically targeting and eliminating a granulosa cell tumor-causing mutation, as well as its therapeutic potential for the treatment of this rare ovarian cancer.
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Project description:FOXL2 is a transcription factor essential for female fertility, expressed in somatic cells of the ovary, notably granulosa cells. In the mouse, Foxl2 deletion leads to partial sex reversal postnatally. However, deletion of the gene in 8-week-old females leads to granulosa to Sertoli cell transdifferentiation. We hypothesised that different outcomes of Foxl2 deletion in embryonic versus adult ovary may depend on a different role played across ovarian development. Therefore, we characterised the dynamics of gene expression and chromatin accessibility changes in purified murine granulosa cells across key developmental stages (E14.5, 1 and 8 weeks). We then performed genome-wide identification of FOXL2 target genes and on-chromatin interacting partners by ChIP-SICAP. We found that FOXL2 regulates more genes at postnatal stages, through the interaction with factors regulating primordial follicle activation (PFA), such as NR5A2, and others regulating steroidogenesis including AR and ESR2. As a proof of principle experiment, we chose one FOXL2 interactor, Ubiquitin specific protease 7 (USP7) and showed that deletion of this gene in granulosa cells leads to a blockage of PFA, impaired ovary development and sterility. Our study constitutes a comprehensive resource for exploration of the molecular mechanisms of ovarian development and causes of female infertility.
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Project description:The Foxl2 transcription factor is required for ovarian function during follicular development. Our approach to begin to understand Foxl2 function is through the identification of Foxl2 regulated genes in the ovary. Transiently transfected KK1 mouse granulosa cells were used to identify genes that are potentially regulated by Foxl2. KK1 cells were transfected in three groups (mock, activated, and repressed) and twenty-four hours later RNA was isolated and submitted for Affymetrix microarray analysis. Experiment Overall Design: To increase the potential of Foxl2 to alter gene expression levels of putative target genes, two fusions were constructed consisting of Foxl2 fused to the activation domain of the Herpes simplex virus VP16 transcription factor (Foxl2-VP16) and Foxl2 fused to the repression domain of the murine MAD transcription factor (Foxl2-MAD). Levels of gene expression were compared between mock transfected cells and those that were transfected with Foxl2-VP16 and Foxl2-MAD, respectively.
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Project description:The identity of the gonads is determined by which fate, ovarian granulosa cell or testicular Sertoli cell, the bipotential somatic cell precursors choose to follow. In most vertebrates, the fate of granulosa cells is controlled by a conserved regulator FOXL2. To understand how FOXL2 elicits its fate-determining action, we performed genome-wide analysis of FOXL2 chromatin occupancy in fetal ovaries. Combining genome-wide analysis of FOXL2 binding in the fetal ovary with transcriptomic analyses of Foxl2 gain-of-function and Foxl2 loss-of-function models, we identified potential pathways responsible for the feminizing action of FOXL2. Finally, comparison of FOXL2 genome-wide occupancy in the fetal ovary with testis-determining factor SOX9 genome-wide occupancy in the fetal testis revealed extensive overlaps, implying that antagonistic signals between FOXL2 and SOX9 occur at the chromatin level.
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