The retinoid-X receptor ortholog, ultraspiracle, binds with nanomolar affinity to an endogenous morphogenetic ligand
The retinoid-X receptor ortholog, ultraspiracle, binds with nanomolar affinity to an endogenous morphogenetic ligand
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DOI：
10.1111/j.1742-4658.2006.05498.x
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发表时间：
2006-11-01
期刊：
影响因子：
5.4
通讯作者：
Wozniak, Mietek
中科院分区：
文献类型：
--
作者：
Jones, Grace;Jones, Davy;Wozniak, Mietek
研究了黑腹果蝇类视黄醇- x受体（RXR）同源物超吸体对环腺天然类法内脂产物的体内配体结合功能和配体结合活性。利用平衡荧光结合法，测定了幼年激素（JH）生物合成途径中的法尼松类产物及其环氧衍生物对超螺旋（USP）的亲和力常数。法尼醇、法尼醇、法尼酸和幼年激素III表现出高纳摩尔到低微摩尔的亲和力，在基本法尼基结构的双键上加入环氧化物后，这种亲和力都降低了。对甲基醚、醇、醛和羧酸在C1上取代的类似分析表明，它们的亲和力都弱于甲酯。因此，将注意力集中在具有甲酯和缺乏环氧化物特征的环腺法内酯产品上。经鉴定，环腺分泌产物法尼酸甲酯具有这些特征，并且对超吸波（K-d = 40 nM）具有与RXR对9-顺式维甲酸相似的亲和力。对侧链延伸到配体结合袋腔内的氨基酸残基的突变分析（不与二级受体结构相互作用，也不延伸到受体表面与共激活剂、辅抑制剂或受体二聚体伴侣相互作用）表明，突变C472A/H475L强烈降低了USP与该环腺产物和JH III的结合。对其他环状腺类激素的影响较小，对JH（非自然的果蝇）结合的影响也很小。与脱皮激素受体一起，USP现在是第二个节肢动物核激素受体，其内分泌腺的分泌产物与纳米摩尔亲和力结合受体已被确定。
The in vivo ligand-binding function and ligand-binding activity of the Drosophila melanogaster retinoid-X receptor (RXR) ortholog, ultraspiracle, toward natural farnesoid products of the ring gland were assessed. Using an equilibrium fluorescence-binding assay, farnesoid products in the juvenile hormone (JH) biosynthesis pathway, and their epoxy derivatives, were measured for their affinity constant for ultraspiracle (USP). Farnesol, farnesal, farnesoic acid and juvenile hormone III exhibited high nanomolar to low micromolar affinity, which in each case decreased upon addition of an epoxide across a double bond of the basic farnesyl structure. Similar analysis of the substitution on C1 of methyl ether, alcohol, aldehyde, and carboxylic acid showed that each conferred weaker affinity than that provided by the methyl ester. Attention was thus focused for a ring-gland farnesoid product that possesses the features of methyl ester and lack of an epoxide. A secreted product of the ring gland, methyl farnesoate, was identified possessing these features and exhibited an affinity for ultraspiracle (K-d = 40 nM) of similar strength to that of RXR for 9-cis retinoic acid. Mutational analysis of amino acid residues with side chains extending into the ligand-binding pocket cavity (and not interacting with secondary receptor structures or extending to the receptor surface to interact with coactivators, corepressors or receptor dimer partners) showed that the mutation C472A/H475L strongly reduced USP binding to this ring gland product and to JH III, with less effect on other ring-gland farnesoids and little effect on binding by (the unnatural to Drosophila) JH I. Along with the ecdysone receptor, USP is now the second arthropod nuclear hormone receptor for which a secreted product of an endocrine gland that binds the receptor with nanomolar affinity has been identified.