The Drosophila Juvenile Hormone Receptor Candidates Methoprene-tolerant (MET) and Germ Cell-expressed (GCE) Utilize a Conserved LIXXL Motif to Bind the FTZ-F1 Nuclear Receptor
The Drosophila Juvenile Hormone Receptor Candidates Methoprene-tolerant (MET) and Germ Cell-expressed (GCE) Utilize a Conserved LIXXL Motif to Bind the FTZ-F1 Nuclear Receptor
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DOI：
10.1074/jbc.m111.327254
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发表时间：
2012-03-02
影响因子：
4.8
通讯作者：
Dubrovsky, Edward B.
中科院分区：
文献类型：
--
作者：
Bernardo, Travis J.;Dubrovsky, Edward B.
保幼激素（JH）参与了全变态昆虫的许多发育过程，但其信号转导机制仍存在争议。我们先前发现，在果蝇Schneider 2细胞中，核受体FTZ-F1是JH激活E75 A基因所必需的。在这里，我们利用昆虫双杂交试验表明，FTZ-F1与两个JH受体候选人，bHLH-PAS旁系同源物MET和GCE，在JH依赖性的方式相互作用。当FTZ-F1激活功能2（AF 2）的螺旋12被去除时，这些相互作用严重减少，暗示AF 2是相互作用位点。通过同源性建模，我们发现MET和GCE具有C-末端α-螺旋，其特征在于代表新的核受体（NR）盒的保守基序LIXXL。由双杂交实验支持的对接模拟显示，FTZ-F1-MET和FTZ-F1-GCE异二聚体的形成涉及典型的NR盒-AF 2相互作用，但不需要FTZ-F1的典型电荷钳残基，并且主要依赖于疏水接触，包括与螺旋4的独特相互作用。此外，我们确定了旁系特异性的功能，包括一个二级相互作用网站只发现在MET。我们的研究结果表明，一种新的NR盒使MET和GCE能够与FTZ-F1的AF 2以JH依赖性相互作用。
Juvenile hormone (JH) has been implicated in many developmental processes in holometabolous insects, but its mechanism of signaling remains controversial. We previously found that in Drosophila Schneider 2 cells, the nuclear receptor FTZ-F1 is required for activation of the E75A gene by JH. Here, we utilized insect two-hybrid assays to show that FTZ-F1 interacts with two JH receptor candidates, the bHLH-PAS paralogs MET and GCE, in a JH-dependent manner. These interactions are severely reduced when helix 12 of the FTZ-F1 activation function 2 (AF2) is removed, implicating AF2 as an interacting site. Through homology modeling, we found that MET and GCE possess a C-terminal alpha-helix featuring a conserved motif LIXXL that represents a novel nuclear receptor (NR) box. Docking simulations supported by two-hybrid experiments revealed that FTZ-F1-MET and FTZ-F1-GCE heterodimer formation involves a typical NR box-AF2 interaction but does not require the canonical charge clamp residues of FTZ-F1 and relies primarily on hydrophobic contacts, including a unique interaction with helix 4. Moreover, we identified paralog-specific features, including a secondary interaction site found only in MET. Our findings suggest that a novel NR box enables MET and GCE to interact JH-dependently with the AF2 of FTZ-F1.