The Amino-terminal Domain of the Androgen Receptor Co-opts Extracellular Signal-regulated Kinase (ERK) Docking Sites in ELK1 Protein to Induce Sustained Gene Activation That Supports Prostate Cancer Cell Growth.
The Amino-terminal Domain of the Androgen Receptor Co-opts Extracellular Signal-regulated Kinase (ERK) Docking Sites in ELK1 Protein to Induce Sustained Gene Activation That Supports Prostate Cancer Cell Growth.
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DOI：
10.1074/jbc.m116.745596
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发表时间：
2016-12-09
期刊：
影响因子：
--
通讯作者：
Ratnam M
中科院分区：
文献类型：
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作者：
Rosati R;Patki M;Chari V;Dakshnamurthy S;McFall T;Saxton J;Kidder BL;Shaw PE;Ratnam M
ETS结构域转录因子ELK 1与生长基因呈抑制性相关，并通过ERK 1/2磷酸化瞬时激活。在前列腺癌（PCa）细胞中，雄激素受体（AR）通过ELK 1的氨基末端结构域（A/B）作为转录共激活因子被ELK 1募集，而没有ELK 1过度磷酸化。在这里，我们阐明了AR与ELK 1相互作用的结构基础。使用哺乳动物双杂交系统对AR共激活所需的ELK 1多肽基序与ERK激活ELK 1所需的ELK 1多肽基序进行了系统性作图，并使用免疫共沉淀试验进行了确认。该定位精确地鉴定了ELK 1中的两个ERK对接位点，即D-box和DEF（ERK对接位点，FXFP）基序，作为其与AR（A/B）或WTAR合作的必要基序。相反，ELK 1中的反式激活结构域仅为ERK激活所需。ELK 1介导的AR（A/B）转录活性在ELK 1结合伴侣、ERK 1/2和血清反应因子不存在时最佳。纯化的ELK 1和AR结合的解离常数为1.9 × 10−8 m。其中D-box和DEF基序被破坏的纯化的突变体ELK 1不结合AR。缺失D-box区域的ELK 1突变体对对MEK抑制不敏感的PCa细胞的雄激素依赖性生长具有显性负效应。这种新的机制，其中一个核受体的信号通路上的冲击，通过增选蛋白激酶对接位点组成性激活生长基因，可以使一类新的靶向药物干预的合理设计。
The ETS domain transcription factor ELK1 is in a repressive association with growth genes and is transiently activated through phosphorylation by ERK1/2. In prostate cancer (PCa) cells the androgen receptor (AR) is recruited by ELK1, via its amino-terminal domain (A/B), as a transcriptional co-activator, without ELK1 hyper-phosphorylation. Here we elucidate the structural basis of the interaction of AR with ELK1. The ELK1 polypeptide motifs required for co-activation by AR versus those required for activation of ELK1 by ERK were systematically mapped using a mammalian two-hybrid system and confirmed using a co-immunoprecipitation assay. The mapping precisely identified the two ERK-docking sites in ELK1, the D-box and the DEF (docking site for ERK, FXFP) motif, as the essential motifs for its cooperation with AR(A/B) or WTAR. In contrast, the transactivation domain in ELK1 was only required for activation by ERK. ELK1-mediated transcriptional activity of AR(A/B) was optimal in the absence of ELK1 binding partners, ERK1/2 and serum-response factor. Purified ELK1 and AR bound with a dissociation constant of 1.9 × 10−8 m. A purified mutant ELK1 in which the D-box and DEF motifs were disrupted did not bind AR. An ELK1 mutant with deletion of the D-box region had a dominant-negative effect on androgen-dependent growth of PCa cells that were insensitive to MEK inhibition. This novel mechanism in which a nuclear receptor impinges on a signaling pathway by co-opting protein kinase docking sites to constitutively activate growth genes could enable rational design of a new class of targeted drug interventions.