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FOXA1 acts upstream of GATA2 and AR in hormonal regulation of gene expression

View Article: PubMed Central - PubMed

ABSTRACT

Hormonal regulation of gene expression by androgen receptor (AR) is tightly controlled by many transcriptional cofactors, including pioneer factors FOXA1 and GATA2, which, however, exhibit distinct expression patterns and functional roles in prostate cancer. Here, we examined how FOXA1, GATA2, and AR crosstalk and regulate hormone-dependent gene expression in prostate cancer cells. ChIP-seq analysis revealed that FOXA1 reprograms both AR and GATA2 cistrome by preferably recruiting them to FKHD-containing genomic sites. By contrast, GATA2 is unable to shift AR or FOXA1 to GATA motifs. Rather, GATA2 co-occupancy enhances AR and FOXA1 binding to nearby ARE and FKHD sites, respectively. Similarly, AR increases, but not re-programs, GATA2 and FOXA1 cistromes. Concordantly, GATA2 and AR strongly enhance the transcriptional program of each other, whereas FOXA1 regulates GATA2- and AR-mediated gene expression in a context-dependent manner due to its reprogramming effects. Taken together, our data delineated for the first time the distinct mechanisms by which GATA2 and FOXA1 regulate AR cistrome and suggest that FOXA1 acts upstream of GATA2 and AR in determining hormone-dependent gene expression in prostate cancer.

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Collaborative effects of GATA2 and AR in transcriptional regulationA–B. Genes induced by GATA2 in androgen-depleted (A) or androgen-stimulated cells (B) are up-regulated by androgen. GATA2-induced gene sets were obtained through microarray analysis of control and GATA2-knockdown LNCaP cells grown in the presence or absence of androgen. GSEA analysis was done in microarray dataset comparing gene expression in LNCaP cells with or without androgen stimulation.C–D. Genes induced by FOXA1 in androgen-depleted LNCaP cells are significantly enriched for down-regulation by androgen (C), while FOXA1-induced genes in the presence of androgen tend to be up-regulated by androgen (D). FOXA1-induced gene sets were obtained through microarray analysis of control and FOXA1-knockdown LNCaP cells grown under androgen-depleted or –replenished conditions and subjected to GSEA analysis.E. Hierarchical clustering showing that GATA2-induced genes in the absence of androgen are further induced by androgen, whereas GATA2-repressed genes are further repressed by androgen. GATA2-induced or –repressed gene sets were derived from microarray data of control and GATA2-knockdown LNCaP cells in the absence of androgen.F. QRT-PCR confirming that GATA2 positively regulates AR activity under both androgen-depleted and –replenished conditions. LNCaP cells grown in the presence or absence of androgen were subjected to control or GATA2 knockdown and then qRT-PCR analysis. PSA and KLK2 are known AR-induced genes, whereas NOV has been reported to be an AR-repressed gene29. Data shown are mean ± SEM in triplicate qPCR and is a representative of at least two independent experiments.
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Figure 3: Collaborative effects of GATA2 and AR in transcriptional regulationA–B. Genes induced by GATA2 in androgen-depleted (A) or androgen-stimulated cells (B) are up-regulated by androgen. GATA2-induced gene sets were obtained through microarray analysis of control and GATA2-knockdown LNCaP cells grown in the presence or absence of androgen. GSEA analysis was done in microarray dataset comparing gene expression in LNCaP cells with or without androgen stimulation.C–D. Genes induced by FOXA1 in androgen-depleted LNCaP cells are significantly enriched for down-regulation by androgen (C), while FOXA1-induced genes in the presence of androgen tend to be up-regulated by androgen (D). FOXA1-induced gene sets were obtained through microarray analysis of control and FOXA1-knockdown LNCaP cells grown under androgen-depleted or –replenished conditions and subjected to GSEA analysis.E. Hierarchical clustering showing that GATA2-induced genes in the absence of androgen are further induced by androgen, whereas GATA2-repressed genes are further repressed by androgen. GATA2-induced or –repressed gene sets were derived from microarray data of control and GATA2-knockdown LNCaP cells in the absence of androgen.F. QRT-PCR confirming that GATA2 positively regulates AR activity under both androgen-depleted and –replenished conditions. LNCaP cells grown in the presence or absence of androgen were subjected to control or GATA2 knockdown and then qRT-PCR analysis. PSA and KLK2 are known AR-induced genes, whereas NOV has been reported to be an AR-repressed gene29. Data shown are mean ± SEM in triplicate qPCR and is a representative of at least two independent experiments.

Mentions: Next, we asked how androgen regulates GATA2 and FOXA1-mediated transcriptional programs. To discern any context-dependent effects, we obtained GATA2-induced and –repressed gene sets through microarray analysis of GATA2-knockdown in the presence and absence of androgen. GSEA revealed that, regardless of the androgen environment, GATA2-induced genes were significantly enriched for up-regulation by androgen (Figure 3A–B), whereas GATA2-repressed genes were further down-regulated following androgen treatment (Figure S3C–D). By contrast, the genes induced by FOXA1 in androgen-depleted LNCaP cells were significantly enriched for down-regulation by androgen (Figure 3C), whereas those induced by FOXA1 in androgen-replenished cells were up-regulated by androgen (Figure 3D), being consistent with its context-dependent roles as a pioneer factor22. Heatmap view of GATA2-induced and –repressed genes further illustrated the collaborative roles of GATA2 and androgen in regulating gene regulation (Figure 3E). Moreover, qRT-PCR analysis confirmed that GATA2 depletion decreased the expression of AR-induced genes such as PSA and KLK2, while restoring AR-repressed gene NOV (Figure 3F). Therefore, FOXA1 and GATA2 play distinct roles in their regulation of AR-mediated transcriptional program; Unlike FOXA1, GATA2 acts as a collaborating transcription factor, rather than a reprogramming factor, of AR.


FOXA1 acts upstream of GATA2 and AR in hormonal regulation of gene expression
Collaborative effects of GATA2 and AR in transcriptional regulationA–B. Genes induced by GATA2 in androgen-depleted (A) or androgen-stimulated cells (B) are up-regulated by androgen. GATA2-induced gene sets were obtained through microarray analysis of control and GATA2-knockdown LNCaP cells grown in the presence or absence of androgen. GSEA analysis was done in microarray dataset comparing gene expression in LNCaP cells with or without androgen stimulation.C–D. Genes induced by FOXA1 in androgen-depleted LNCaP cells are significantly enriched for down-regulation by androgen (C), while FOXA1-induced genes in the presence of androgen tend to be up-regulated by androgen (D). FOXA1-induced gene sets were obtained through microarray analysis of control and FOXA1-knockdown LNCaP cells grown under androgen-depleted or –replenished conditions and subjected to GSEA analysis.E. Hierarchical clustering showing that GATA2-induced genes in the absence of androgen are further induced by androgen, whereas GATA2-repressed genes are further repressed by androgen. GATA2-induced or –repressed gene sets were derived from microarray data of control and GATA2-knockdown LNCaP cells in the absence of androgen.F. QRT-PCR confirming that GATA2 positively regulates AR activity under both androgen-depleted and –replenished conditions. LNCaP cells grown in the presence or absence of androgen were subjected to control or GATA2 knockdown and then qRT-PCR analysis. PSA and KLK2 are known AR-induced genes, whereas NOV has been reported to be an AR-repressed gene29. Data shown are mean ± SEM in triplicate qPCR and is a representative of at least two independent experiments.
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Related In: Results  -  Collection

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Figure 3: Collaborative effects of GATA2 and AR in transcriptional regulationA–B. Genes induced by GATA2 in androgen-depleted (A) or androgen-stimulated cells (B) are up-regulated by androgen. GATA2-induced gene sets were obtained through microarray analysis of control and GATA2-knockdown LNCaP cells grown in the presence or absence of androgen. GSEA analysis was done in microarray dataset comparing gene expression in LNCaP cells with or without androgen stimulation.C–D. Genes induced by FOXA1 in androgen-depleted LNCaP cells are significantly enriched for down-regulation by androgen (C), while FOXA1-induced genes in the presence of androgen tend to be up-regulated by androgen (D). FOXA1-induced gene sets were obtained through microarray analysis of control and FOXA1-knockdown LNCaP cells grown under androgen-depleted or –replenished conditions and subjected to GSEA analysis.E. Hierarchical clustering showing that GATA2-induced genes in the absence of androgen are further induced by androgen, whereas GATA2-repressed genes are further repressed by androgen. GATA2-induced or –repressed gene sets were derived from microarray data of control and GATA2-knockdown LNCaP cells in the absence of androgen.F. QRT-PCR confirming that GATA2 positively regulates AR activity under both androgen-depleted and –replenished conditions. LNCaP cells grown in the presence or absence of androgen were subjected to control or GATA2 knockdown and then qRT-PCR analysis. PSA and KLK2 are known AR-induced genes, whereas NOV has been reported to be an AR-repressed gene29. Data shown are mean ± SEM in triplicate qPCR and is a representative of at least two independent experiments.
Mentions: Next, we asked how androgen regulates GATA2 and FOXA1-mediated transcriptional programs. To discern any context-dependent effects, we obtained GATA2-induced and –repressed gene sets through microarray analysis of GATA2-knockdown in the presence and absence of androgen. GSEA revealed that, regardless of the androgen environment, GATA2-induced genes were significantly enriched for up-regulation by androgen (Figure 3A–B), whereas GATA2-repressed genes were further down-regulated following androgen treatment (Figure S3C–D). By contrast, the genes induced by FOXA1 in androgen-depleted LNCaP cells were significantly enriched for down-regulation by androgen (Figure 3C), whereas those induced by FOXA1 in androgen-replenished cells were up-regulated by androgen (Figure 3D), being consistent with its context-dependent roles as a pioneer factor22. Heatmap view of GATA2-induced and –repressed genes further illustrated the collaborative roles of GATA2 and androgen in regulating gene regulation (Figure 3E). Moreover, qRT-PCR analysis confirmed that GATA2 depletion decreased the expression of AR-induced genes such as PSA and KLK2, while restoring AR-repressed gene NOV (Figure 3F). Therefore, FOXA1 and GATA2 play distinct roles in their regulation of AR-mediated transcriptional program; Unlike FOXA1, GATA2 acts as a collaborating transcription factor, rather than a reprogramming factor, of AR.

View Article: PubMed Central - PubMed

ABSTRACT

Hormonal regulation of gene expression by androgen receptor (AR) is tightly controlled by many transcriptional cofactors, including pioneer factors FOXA1 and GATA2, which, however, exhibit distinct expression patterns and functional roles in prostate cancer. Here, we examined how FOXA1, GATA2, and AR crosstalk and regulate hormone-dependent gene expression in prostate cancer cells. ChIP-seq analysis revealed that FOXA1 reprograms both AR and GATA2 cistrome by preferably recruiting them to FKHD-containing genomic sites. By contrast, GATA2 is unable to shift AR or FOXA1 to GATA motifs. Rather, GATA2 co-occupancy enhances AR and FOXA1 binding to nearby ARE and FKHD sites, respectively. Similarly, AR increases, but not re-programs, GATA2 and FOXA1 cistromes. Concordantly, GATA2 and AR strongly enhance the transcriptional program of each other, whereas FOXA1 regulates GATA2- and AR-mediated gene expression in a context-dependent manner due to its reprogramming effects. Taken together, our data delineated for the first time the distinct mechanisms by which GATA2 and FOXA1 regulate AR cistrome and suggest that FOXA1 acts upstream of GATA2 and AR in determining hormone-dependent gene expression in prostate cancer.

No MeSH data available.


Related in: MedlinePlus