Laboratories for Reproductive Biology, Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC 27599-7500, USA.
Mol Cell Endocrinol. 2012 Jan 30;348(2):403-10. doi: 10.1016/j.mce.2011.03.026. Epub 2011 Jun 1.
Human androgen receptor (AR) transcriptional activity involves interdomain and coactivator interactions with the agonist-bound AR ligand binding domain (LBD). Structural determinants of the AR NH(2)- and carboxyl-terminal interaction between the AR NH(2)-terminal FXXLF motif and activation function 2 (AF2) in the LBD were shown previously by crystallography. In this report, we provide evidence for a region in AR LBD helix 12 outside the AF2 binding cleft that facilitates interactions with the FXXLF and LXXLL motifs. Mutagenesis of glutamine 902 to alanine in AR LBD helix 12 (Q902A) disrupted AR FXXLF motif binding to AF2, but enhanced coactivator LXXLL motif binding. Functional compensation for defective FXXLF motif binding by AR-Q902A was suggested by the slower dissociation rate of bound androgen. Functional importance of glutamine 902 was indicated by the charged residue germline mutation Q902R that caused partial androgen insensitivity, and a similar somatic mutation Q902K reported in prostate cancer, both of which increased the androgen dissociation rate and decreased AR transcriptional activity. High affinity equilibrium androgen binding was retained by alanine substitution mutations at Tyr-739 in AR LBD helix 5 or Lys-905 in helix 12 structurally adjacent to AF2, whereas transcriptional activity decreased and the androgen dissociation increased. Deleterious effects of these loss of function mutations were rescued by the helix stabilizing AR prostate cancer somatic mutation H874Y. Sequence NH(2)-terminal to the AR FXXLF motif contributed to the AR NH(2)- and carboxyl-terminal interaction based on greater AR-2-30 FXXLF motif peptide binding to the agonist-bound AR LBD than a shorter AR-20-30 FXXLF motif peptide. We conclude that helix 12 residues outside the AF2 binding cleft modulate AR transcriptional activity by providing flexibility to accommodate FXXLF or LXXLL motif binding.
人类雄激素受体 (AR) 的转录活性涉及与激动剂结合的 AR 配体结合域 (LBD) 之间的结构域间和共激活因子相互作用。先前通过晶体学研究显示了 AR NH(2)-末端的 FXXLF 基序和 LBD 中的激活功能 2 (AF2) 之间的 AR NH(2)-末端和羧基末端相互作用的结构决定因素。在本报告中，我们提供了证据表明 AR LBD 螺旋 12 中 AF2 结合裂隙之外的一个区域有助于与 FXXLF 和 LXXLL 基序相互作用。AR LBD 螺旋 12 中的谷氨酰胺 902 突变为丙氨酸 (Q902A) 破坏了 AR FXXLF 基序与 AF2 的结合，但增强了共激活因子 LXXLL 基序的结合。AR-Q902A 结合缺陷的 FXXLF 基序的功能补偿表明，结合的雄激素的解离速率较慢。通过 Q902R 这种带电荷的种系突变引起部分雄激素不敏感，以及在前列腺癌中报道的类似的体细胞突变 Q902K，都表明谷氨酰胺 902 的重要性，这两种突变都增加了雄激素的解离速率并降低了 AR 转录活性。在与 AF2 结构相邻的 AR LBD 螺旋 5 中的 Tyr-739 或螺旋 12 中的 Lys-905 处用丙氨酸取代突变时，保留了高亲和力平衡雄激素结合，而转录活性降低，雄激素解离增加。这些功能丧失突变的有害影响可以通过 AR 前列腺癌体细胞突变 H874Y 来挽救。AR FXXLF 基序的 NH(2)-末端序列基于 AR-2-30 FXXLF 基序肽与激动剂结合的 AR LBD 的结合比更短的 AR-20-30 FXXLF 基序肽更强，对 AR NH(2)-和羧基末端相互作用有贡献。我们得出结论，AF2 结合裂隙之外的螺旋 12 残基通过提供灵活性来适应 FXXLF 或 LXXLL 基序的结合，从而调节 AR 转录活性。
