A Role of the Amino-Terminal (N) and Carboxyl-Terminal (C) Interaction in Binding of

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A Role of the Amino-Terminal (N) and Carboxyl-Terminal (C) Interaction in Binding of Androgen Receptor to Chromatin
Jiwen Li, Junjiang Fu, Charalambos Toumazou, Ho-Geun Yoon, Jiemin Wong
Molecular Endocrinology, Volume 20, Issue 4, 1 April 2006, Pages 776–785, https://doi.org/10.1210/me.2005-0298
Published:
01 April 2006
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Abstract

The N-terminal domain of AR is known to engage a hormone-dependent interaction with its C-terminal ligand-binding domain, and this N/C interaction is known to modulate AR transcriptional activity. Using Xenopus oocytes as a model system to study transcriptional regulation in chromatin, we found that two previously reported N/C interaction-defective AR mutants, one with deletion of 23FQNLF27(ARΔF) and one with a Gly 21 to Glu mutation (ARG21E), were surprisingly inactive in activating transcription from various reporters assembled into chromatin. Further study using chromatin immunoprecipitation assay revealed that these mutants failed to bind both mouse mammary tumor virus-long terminal repeat and prostate-specific antigen enhancer assembled into chromatin. This defect is specific to chromatin because both mutants could bind to a consensus AR response element in vitro and activate transcription driven by mouse mammary tumor virus-long terminal repeat in transient transfection as effective as the wild-type AR. To further substantiate this novel finding, we established 293 cell lines that stably expressed either AR or ARΔF mutant in an inducible manner. Using these cell lines, we confirmed by using chromatin immunoprecipitation assay that AR but not ARΔF could bind to the endogenous prostate-specific antigen enhancer. Furthermore, we found that the ARΔF mutant interacts poorly with Brg1, the ATPase subunit of the chromatin-remodeling factor SWI/SNF. Taken together, our study reveals a novel role of AR N/C interaction in control of AR chromatin binding and suggests a working model that the proper N/C interaction is required for AR to recruit SWI/SNF complex, which in turn remodels chromatin to allow AR to bind to AR response elements in chromatin.

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Analysis of Interdomain Interactions of the Androgen Receptor
Elizabeth M. Wilson
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Abstract

High-affinity binding of testosterone or dihydrotestosterone to the androgen receptor (AR) triggers the androgen-dependent AR NH2- and carboxyl-terminal (N/C) interaction between the AR NH2-terminal FXXLF motif and the activation function 2 (AF2) hydrophobic binding surface in the ligand-binding domain. The functional importance of the AR N/C interaction is supported by naturally occurring loss-of-function AR AF2 mutations where AR retains high-affinity androgen binding but is defective in AR FXXLF motif binding. Ligands with agonist activity in vivo such as testosterone, dihydrotestosterone, and the synthetic anabolic steroids induce the AR N/C interaction and increase AR transcriptional activity in part by slowing the dissociation rate of bound ligand and stabilizing AR against degradation. AR ligand-binding domain competitive antagonists inhibit the agonist-dependent AR N/C interaction. Although the human AR N/C interaction is important for transcriptional activity, it has an inhibitory effect on transcriptional activity from AF2 by competing for p160 coactivator LXXLL motif binding. The primate-specific AR coregulatory protein, melanoma antigen gene protein-A11 (MAGE-A11), modulates the AR N/C interaction through a direct interaction with the AR FXXLF motif. Inhibition of AF2 transcriptional activity by the AR N/C interaction is relieved by AR FXXLF motif binding to the F-box region of MAGE-11. Described here are methods to measure the androgen-dependent AR N/C inter-domain interaction and the influence of transcriptional coregulators.
Keywords: Androgen receptor, steroid receptor, N/C interaction, mammalian two-hybrid assay, MAGE-11, MAGE-A11