Limits...
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.

No MeSH data available.


Related in: MedlinePlus

AR co-occupancy potentiates GATA2 binding on the chromatinA. Western Blot analysis of GATA2 and AR in LNCaP cells in the presence (FBS) and absence (FCS) of androgen.B. Venn diagram showing overlap between GATA binding sites in androgen-depleted and androgen-stimulated LNCaP cells.C. Motif analysis of ARE, GATA and FKHD enrichment in the three categories of GTBS as shown in B.D. Heatmap of AR and GATA2 ChIP-seq read intensity around the three categories of GTBS identified in B.E. Genome browser view showing increased GATA2 binding at the TMPRSS2 gene enhancer in androgen-stimulated cells.F. ChIP-PCR showing GATA2 enrichment at the TMPRSS2 and PSA enhancers in LNCaP cells in the absence (FCS) or presence of androgen (FBS). Data shown are mean ± SEM in triplicate qPCR and is a representative of at least two independent experiments.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC4940300&req=5

Figure 2: AR co-occupancy potentiates GATA2 binding on the chromatinA. Western Blot analysis of GATA2 and AR in LNCaP cells in the presence (FBS) and absence (FCS) of androgen.B. Venn diagram showing overlap between GATA binding sites in androgen-depleted and androgen-stimulated LNCaP cells.C. Motif analysis of ARE, GATA and FKHD enrichment in the three categories of GTBS as shown in B.D. Heatmap of AR and GATA2 ChIP-seq read intensity around the three categories of GTBS identified in B.E. Genome browser view showing increased GATA2 binding at the TMPRSS2 gene enhancer in androgen-stimulated cells.F. ChIP-PCR showing GATA2 enrichment at the TMPRSS2 and PSA enhancers in LNCaP cells in the absence (FCS) or presence of androgen (FBS). Data shown are mean ± SEM in triplicate qPCR and is a representative of at least two independent experiments.

Mentions: As GATA2-mediated transcriptional regulation has recently been shown to play important roles in prostate tumorigenesis20, we next asked whether AR might also enhance GATA2 function, forming a potential feed-forward loop further contributing to prostate cancer progression. We first performed western blotting of LNCaP cells grown in the presence (FBS) or absence of androgen (FCS). Our data demonstrated that, as previously reported21, androgen stimulation drastically increased AR protein level in LNCaP cells. Interestingly, GATA2 protein level was slightly decreased in the presence of androgen (Figure 2A). ChIP-seq analysis showed that, as expected, there are many more AR binding events in the presence of androgen (Figure S2A). Importantly, despite the significant decrease of GATA2 protein, ChIP-seq analysis revealed an increase in total number of GTBS and a clear shift of GTBS to ARE-enriched regions upon androgen stimulation (Figure 2B–C). This suggests that AR occupancy on the chromatin might facilitate GATA2 binding to the same sites. Indeed, heatmap view of ChIP-seq data demonstrated that AR is much more enriched at conserved (category II) and gained GTBS (III) and AR enrichment were further enhanced under FBS conditions (Figure 2D).


FOXA1 acts upstream of GATA2 and AR in hormonal regulation of gene expression
AR co-occupancy potentiates GATA2 binding on the chromatinA. Western Blot analysis of GATA2 and AR in LNCaP cells in the presence (FBS) and absence (FCS) of androgen.B. Venn diagram showing overlap between GATA binding sites in androgen-depleted and androgen-stimulated LNCaP cells.C. Motif analysis of ARE, GATA and FKHD enrichment in the three categories of GTBS as shown in B.D. Heatmap of AR and GATA2 ChIP-seq read intensity around the three categories of GTBS identified in B.E. Genome browser view showing increased GATA2 binding at the TMPRSS2 gene enhancer in androgen-stimulated cells.F. ChIP-PCR showing GATA2 enrichment at the TMPRSS2 and PSA enhancers in LNCaP cells in the absence (FCS) or presence of androgen (FBS). Data shown are mean ± SEM in triplicate qPCR and is a representative of at least two independent experiments.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4940300&req=5

Figure 2: AR co-occupancy potentiates GATA2 binding on the chromatinA. Western Blot analysis of GATA2 and AR in LNCaP cells in the presence (FBS) and absence (FCS) of androgen.B. Venn diagram showing overlap between GATA binding sites in androgen-depleted and androgen-stimulated LNCaP cells.C. Motif analysis of ARE, GATA and FKHD enrichment in the three categories of GTBS as shown in B.D. Heatmap of AR and GATA2 ChIP-seq read intensity around the three categories of GTBS identified in B.E. Genome browser view showing increased GATA2 binding at the TMPRSS2 gene enhancer in androgen-stimulated cells.F. ChIP-PCR showing GATA2 enrichment at the TMPRSS2 and PSA enhancers in LNCaP cells in the absence (FCS) or presence of androgen (FBS). Data shown are mean ± SEM in triplicate qPCR and is a representative of at least two independent experiments.
Mentions: As GATA2-mediated transcriptional regulation has recently been shown to play important roles in prostate tumorigenesis20, we next asked whether AR might also enhance GATA2 function, forming a potential feed-forward loop further contributing to prostate cancer progression. We first performed western blotting of LNCaP cells grown in the presence (FBS) or absence of androgen (FCS). Our data demonstrated that, as previously reported21, androgen stimulation drastically increased AR protein level in LNCaP cells. Interestingly, GATA2 protein level was slightly decreased in the presence of androgen (Figure 2A). ChIP-seq analysis showed that, as expected, there are many more AR binding events in the presence of androgen (Figure S2A). Importantly, despite the significant decrease of GATA2 protein, ChIP-seq analysis revealed an increase in total number of GTBS and a clear shift of GTBS to ARE-enriched regions upon androgen stimulation (Figure 2B–C). This suggests that AR occupancy on the chromatin might facilitate GATA2 binding to the same sites. Indeed, heatmap view of ChIP-seq data demonstrated that AR is much more enriched at conserved (category II) and gained GTBS (III) and AR enrichment were further enhanced under FBS conditions (Figure 2D).

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