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CCAR1 promotes chromatin loading of androgen receptor (AR) transcription complex by stabilizing the association between AR and GATA2.

Seo WY, Jeong BC, Yu EJ, Kim HJ, Kim SH, Lim JE, Kwon GY, Lee HM, Kim JH - Nucleic Acids Res. (2013)

Bottom Line: Depletion of CCAR1 caused reduction in androgen-dependent expression of a subset of AR target genes.The molecular mechanism underlying CCAR1 function in AR-mediated transcription involves CCAR1-mediated enhanced recruitment of GATA2, a pioneer factor for AR, to AR-binding sites.CCAR1 stabilized the interaction between AR and GATA2 by interacting directly with both proteins, thereby facilitating AR and GATA2 occupancy on the enhancers.

View Article: PubMed Central - PubMed

Affiliation: Department of Health Sciences and Technology, Samsung Advanced Institute for Health Sciences and Technology, Sungkyunkwan University, Seoul, 135-710, Korea, Department of Biomedical Sciences, Samsung Biomedical Research Institute, Samsung Medical Center, Seoul, 135-710, Korea, Department of Urology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, 135-710, Korea and Department of Pathology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, 135-710, Korea.

ABSTRACT
Androgen receptor (AR), a ligand-dependent transcription factor, plays a critical role in prostate cancer onset and progression, and its transcriptional function is mediated largely by distinct nuclear receptor co-regulators. Here, we show that cell cycle and apoptosis regulator 1 (CCAR1) functions as an AR co-activator. CCAR1 interacted with and enhanced the transcriptional activity of AR. Depletion of CCAR1 caused reduction in androgen-dependent expression of a subset of AR target genes. We further showed that CCAR1 is required for recruitment of AR, MED1 and RNA polymerase II to the enhancers of AR target genes and for androgen-induced long-range prostate specific antigen enhancer-promoter interaction. The molecular mechanism underlying CCAR1 function in AR-mediated transcription involves CCAR1-mediated enhanced recruitment of GATA2, a pioneer factor for AR, to AR-binding sites. CCAR1 stabilized the interaction between AR and GATA2 by interacting directly with both proteins, thereby facilitating AR and GATA2 occupancy on the enhancers. Furthermore, CCAR1 depletion inhibited the growth, migration, invasion of prostate cancer cells and reduced the tumorigenicity of prostate cancer cells in vivo. Our results firmly established CCAR1 as an AR co-activator that plays a key role in AR transcription complex assembly and has an important physiological role in androgen signaling and prostate tumorigenesis.

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The proposed role of CCAR1 in chromatin loading and assembly of AR transcription complex. CCAR1 is recruited to GATA2-dependent AR-binding sites at least in part through interaction with DNA-bound GATA2 and AR and stabilizes their association with DNA, thereby increasing their occupancy on AR-binding sites. The recruited CCAR1 possibly provides a physical link between DNA-bound AR and Mediator complex and facilitates recruitment of p160 co-activator complex, and thereby facilitating transcription complex assembly, chromatin looping and chromatin remodeling. For example, CCAR1 facilitates AR and GATA2 recruitment and stabilizes their occupancies on the distal PSA enhancer (ARE III) containing a GATA2-dependent ARE. CCAR1 also facilitates recruitment of Mediator complex and Pol II to AR-bound PSA enhancer (ARE III) and promoter (ARE I and ARE II). CCAR1 provides a physical link in AR transcription complex, thereby leading to chromatin looping and enhancing communication between the PSA enhancer and promoter.
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gkt644-F6: The proposed role of CCAR1 in chromatin loading and assembly of AR transcription complex. CCAR1 is recruited to GATA2-dependent AR-binding sites at least in part through interaction with DNA-bound GATA2 and AR and stabilizes their association with DNA, thereby increasing their occupancy on AR-binding sites. The recruited CCAR1 possibly provides a physical link between DNA-bound AR and Mediator complex and facilitates recruitment of p160 co-activator complex, and thereby facilitating transcription complex assembly, chromatin looping and chromatin remodeling. For example, CCAR1 facilitates AR and GATA2 recruitment and stabilizes their occupancies on the distal PSA enhancer (ARE III) containing a GATA2-dependent ARE. CCAR1 also facilitates recruitment of Mediator complex and Pol II to AR-bound PSA enhancer (ARE III) and promoter (ARE I and ARE II). CCAR1 provides a physical link in AR transcription complex, thereby leading to chromatin looping and enhancing communication between the PSA enhancer and promoter.

Mentions: Genome-wide ChIP-on-chip and ChIP-Seq studies identified a highly significant overlap of AR and GATA2-binding sites in androgen target genes and showed that GATA2 acts as a pioneer factor in the recruitment of AR to AR binding sites (5,6,17). In this study, we report a novel mechanism for AR-mediated transcription involving the interplay between AR, GATA2 and CCAR1. Our ChIP analyses showed that recruitment of AR and GATA2 to the PSA, TMPRSS2 and KLK2 enhancers was decreased by CCAR1 depletion (Figure 4B and C and Supplementary Figure S11A). In line with these observations, MED1 and Pol II failed to bind to the enhancers, and consequently, the expression of PSA, TMPRSS2 and KLK2 was significantly reduced (Figure 2B and Supplementary Figure S4). These results suggest that AR and GATA2 bind to the enhancer more efficiently when complexed with CCAR1. Indeed, CCAR1 stabilized the interaction between AR and GATA2 by associating with both proteins (Figure 5C) and enhanced their DNA-binding activity (Figure 5D). Although GATA2 is required for PSA, TMPRSS2 and KLK2 expression (5,18), GATA2 does not play a significant role in androgen-regulated FKBP5 expression (19). Interestingly, CCAR1 depletion did not affect AR binding but greatly reduced MED1 and Pol II loading on the FKBP5 enhancer (Supplementary Figure S11B), which is similar to our previous finding that inhibition of CCAR1 recruitment to the pS2 promoter had no observable effect on recruitment of ERα but compromised the hormone-dependent recruitment of Mediator complex and Pol II to the promoter (8). Similarly, CCAR1 depletion caused only a modest reduction in AR recruitment to the PSA promoter containing GATA2-independent AREs (Supplementary Figure S12). Thus, we propose that CCAR1 has a dual function in regulating AR signaling: first as an adaptor protein by stabilizing the association between AR and GATA2 and increasing their occupancy on AR-binding sites, and second as a co-activator by facilitating recruitment of Mediator complex and Pol II to AR-binding sites by providing a physical link between DNA-bound AR and the Mediator complex, and thereby facilitating transcription complex assembly, chromatin looping and chromatin accessibility (Figure 6).Figure 6.


CCAR1 promotes chromatin loading of androgen receptor (AR) transcription complex by stabilizing the association between AR and GATA2.

Seo WY, Jeong BC, Yu EJ, Kim HJ, Kim SH, Lim JE, Kwon GY, Lee HM, Kim JH - Nucleic Acids Res. (2013)

The proposed role of CCAR1 in chromatin loading and assembly of AR transcription complex. CCAR1 is recruited to GATA2-dependent AR-binding sites at least in part through interaction with DNA-bound GATA2 and AR and stabilizes their association with DNA, thereby increasing their occupancy on AR-binding sites. The recruited CCAR1 possibly provides a physical link between DNA-bound AR and Mediator complex and facilitates recruitment of p160 co-activator complex, and thereby facilitating transcription complex assembly, chromatin looping and chromatin remodeling. For example, CCAR1 facilitates AR and GATA2 recruitment and stabilizes their occupancies on the distal PSA enhancer (ARE III) containing a GATA2-dependent ARE. CCAR1 also facilitates recruitment of Mediator complex and Pol II to AR-bound PSA enhancer (ARE III) and promoter (ARE I and ARE II). CCAR1 provides a physical link in AR transcription complex, thereby leading to chromatin looping and enhancing communication between the PSA enhancer and promoter.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3794601&req=5

gkt644-F6: The proposed role of CCAR1 in chromatin loading and assembly of AR transcription complex. CCAR1 is recruited to GATA2-dependent AR-binding sites at least in part through interaction with DNA-bound GATA2 and AR and stabilizes their association with DNA, thereby increasing their occupancy on AR-binding sites. The recruited CCAR1 possibly provides a physical link between DNA-bound AR and Mediator complex and facilitates recruitment of p160 co-activator complex, and thereby facilitating transcription complex assembly, chromatin looping and chromatin remodeling. For example, CCAR1 facilitates AR and GATA2 recruitment and stabilizes their occupancies on the distal PSA enhancer (ARE III) containing a GATA2-dependent ARE. CCAR1 also facilitates recruitment of Mediator complex and Pol II to AR-bound PSA enhancer (ARE III) and promoter (ARE I and ARE II). CCAR1 provides a physical link in AR transcription complex, thereby leading to chromatin looping and enhancing communication between the PSA enhancer and promoter.
Mentions: Genome-wide ChIP-on-chip and ChIP-Seq studies identified a highly significant overlap of AR and GATA2-binding sites in androgen target genes and showed that GATA2 acts as a pioneer factor in the recruitment of AR to AR binding sites (5,6,17). In this study, we report a novel mechanism for AR-mediated transcription involving the interplay between AR, GATA2 and CCAR1. Our ChIP analyses showed that recruitment of AR and GATA2 to the PSA, TMPRSS2 and KLK2 enhancers was decreased by CCAR1 depletion (Figure 4B and C and Supplementary Figure S11A). In line with these observations, MED1 and Pol II failed to bind to the enhancers, and consequently, the expression of PSA, TMPRSS2 and KLK2 was significantly reduced (Figure 2B and Supplementary Figure S4). These results suggest that AR and GATA2 bind to the enhancer more efficiently when complexed with CCAR1. Indeed, CCAR1 stabilized the interaction between AR and GATA2 by associating with both proteins (Figure 5C) and enhanced their DNA-binding activity (Figure 5D). Although GATA2 is required for PSA, TMPRSS2 and KLK2 expression (5,18), GATA2 does not play a significant role in androgen-regulated FKBP5 expression (19). Interestingly, CCAR1 depletion did not affect AR binding but greatly reduced MED1 and Pol II loading on the FKBP5 enhancer (Supplementary Figure S11B), which is similar to our previous finding that inhibition of CCAR1 recruitment to the pS2 promoter had no observable effect on recruitment of ERα but compromised the hormone-dependent recruitment of Mediator complex and Pol II to the promoter (8). Similarly, CCAR1 depletion caused only a modest reduction in AR recruitment to the PSA promoter containing GATA2-independent AREs (Supplementary Figure S12). Thus, we propose that CCAR1 has a dual function in regulating AR signaling: first as an adaptor protein by stabilizing the association between AR and GATA2 and increasing their occupancy on AR-binding sites, and second as a co-activator by facilitating recruitment of Mediator complex and Pol II to AR-binding sites by providing a physical link between DNA-bound AR and the Mediator complex, and thereby facilitating transcription complex assembly, chromatin looping and chromatin accessibility (Figure 6).Figure 6.

Bottom Line: Depletion of CCAR1 caused reduction in androgen-dependent expression of a subset of AR target genes.The molecular mechanism underlying CCAR1 function in AR-mediated transcription involves CCAR1-mediated enhanced recruitment of GATA2, a pioneer factor for AR, to AR-binding sites.CCAR1 stabilized the interaction between AR and GATA2 by interacting directly with both proteins, thereby facilitating AR and GATA2 occupancy on the enhancers.

View Article: PubMed Central - PubMed

Affiliation: Department of Health Sciences and Technology, Samsung Advanced Institute for Health Sciences and Technology, Sungkyunkwan University, Seoul, 135-710, Korea, Department of Biomedical Sciences, Samsung Biomedical Research Institute, Samsung Medical Center, Seoul, 135-710, Korea, Department of Urology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, 135-710, Korea and Department of Pathology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, 135-710, Korea.

ABSTRACT
Androgen receptor (AR), a ligand-dependent transcription factor, plays a critical role in prostate cancer onset and progression, and its transcriptional function is mediated largely by distinct nuclear receptor co-regulators. Here, we show that cell cycle and apoptosis regulator 1 (CCAR1) functions as an AR co-activator. CCAR1 interacted with and enhanced the transcriptional activity of AR. Depletion of CCAR1 caused reduction in androgen-dependent expression of a subset of AR target genes. We further showed that CCAR1 is required for recruitment of AR, MED1 and RNA polymerase II to the enhancers of AR target genes and for androgen-induced long-range prostate specific antigen enhancer-promoter interaction. The molecular mechanism underlying CCAR1 function in AR-mediated transcription involves CCAR1-mediated enhanced recruitment of GATA2, a pioneer factor for AR, to AR-binding sites. CCAR1 stabilized the interaction between AR and GATA2 by interacting directly with both proteins, thereby facilitating AR and GATA2 occupancy on the enhancers. Furthermore, CCAR1 depletion inhibited the growth, migration, invasion of prostate cancer cells and reduced the tumorigenicity of prostate cancer cells in vivo. Our results firmly established CCAR1 as an AR co-activator that plays a key role in AR transcription complex assembly and has an important physiological role in androgen signaling and prostate tumorigenesis.

Show MeSH
Related in: MedlinePlus