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Control of androgen receptor action by a novel nuclear receptor binding motif in Bag-1L

  • 1, 2,
  • 3,
  • 3,
  • 3 and
  • 1, 2
Epigenetics & Chromatin20136 (Suppl 1) :P13

https://doi.org/10.1186/1756-8935-6-S1-P13

  • Published:

Keywords

  • Androgen Receptor
  • Cell Line LNCaP
  • Androgen Receptor Binding
  • Prostate Cancer Cell Line LNCaP
  • Cancer Cell Line LNCaP

Background

The androgen receptor (AR) is an important determinant of normal and malignant prostate growth. Therefore, a good understanding of the factors that regulate the transactivation of the AR is essential and could provide a better strategy to control prostate tumor growth, particularly in patients suffering from the hormone-refractory and the advanced stages of the disease. However, transcriptional activation by the AR is a complex and orchestrated process requiring multiple coregulators. Several coregulators have already been identified, including proteasome components, chromatin-re-modeling complexes and heat shock proteins. Others however remain uncharacterized and their role in AR transactivation is poorly understood. One such coregulator is the nuclear-resident, AR co-activator, Bag-1L. Overexpression of Bag-1L and the amplification of its gene have been reported in the hormone-refractory and metastatic stages of prostate cancer and in androgen-independent prostate cancer cells (AIPC). However the exact mechanism of Bag-1L-mediated regulation of AR action in prostate cancer is unclear.

Materials and methods

To confirm the contribution of Bag-1L to AR response in prostate cancer, we have downregulated the expression of Bag-1L in the androgen-dependent prostate cancer cell line LNCaP using RNAi. Subsequently we performed genome-wide mapping of Bag-1L, using genome-wide chromatin immunoprecipitation (ChIP-seq). Lastly, the ability of AR and Bag-1L to interact with one another was assessed by domain mapping experiments using GST pulldown assays.

Results

Here we show that Bag-1L depletion by RNA interference in the prostate cell line LNCaP significantly reduces the hormone response of several AR-target genes. In agreement, Bag-1L and AR co-localize to a large number of regulatory regions of AR-target genes as identified by ChIP-sequencing. Furthermore, domain mapping experiments identified the first 128 N-terminal amino acids of Bag-1L as necessary for enhancing the transactivation function of the receptor. This sequence contains a duplication of a GARRPR motif, which we identified as the main interaction site between Bag-1L and AR. We were able to further confirm the importance of this motif by amino acid substitutions of the sequence, which impairs the AR and Bag-1L-mediated AR transactivation. Intriguingly, overexpression of the N-terminal sequence of Bag-1L encompassing the conserved hexapeptide sequences exerts a dominant negative effect on androgen-mediated gene expression and androgen-dependent tumor growth. The GARRPR was also identified in other regulators of AR activity, such as Huntington associated protein 1, nuclear receptor coactivator 4 and p21-activated kinase 6.

Conclusions

We have identified a novel AR binding motif different from the previously described LXXLL and FXXLF sequences of other coactivators of AR. In the long-term we propose to use this knowledge to design novel drugs targeting this interaction site for the therapeutic intervention of prostate cancer.

Authors’ Affiliations

(1)
Department of Medical Oncology, Dana-Farber Cancer Institute and Harvard Medical School, Boston, USA
(2)
Center for Functional Cancer Epigenetics, Dana-Farber Cancer Institute, Boston, USA
(3)
Institute of Toxicology and Genetics, Karlsruhe Institute of Technology KIT, Eggenstein-Leopoldshafen, Germany

Copyright

© Cato et al; licensee BioMed Central Ltd. 2013

This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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