Dissecting molecular differences between Wnt coreceptors LRP5 and LRP6.
Dissecting molecular differences between Wnt coreceptors LRP5 and LRP6.
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DOI：
10.1371/journal.pone.0023537
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发表时间：
2011
期刊：
影响因子：
3.7
通讯作者：
He X
中科院分区：
文献类型：
--
作者：
MacDonald BT;Semenov MV;Huang H;He X
低密度脂蛋白受体相关蛋白5和6（LRP 5和LRP 6）作为经典β-连环蛋白途径的Wnt共受体。虽然LRP 6对胚胎发育至关重要，但LRP 5和LRP 6在骨骼重塑、骨质疏松症发病机制和癌症形成中发挥关键作用，使得LRP 5和LRP 6成为癌症和疾病治疗的关键治疗靶点。LRP 5和LRP 6在胞质结构域中各自含有5个保守的PPPSPxS基序，其对于信号传导是关键的，并且共同充当支架蛋白Axin的磷酸化依赖性对接位点。然而，现有数据表明LRP 6在转导Wnt信号方面比LRP 5更有效。为了理解LRP 5和LRP 6不同信号传导活性的分子基础，我们通过交换LRP 5和LRP 6胞质结构域，LRP 5C和LRP 6C，产生了一系列嵌合受体，并使用生物化学和功能测定研究了它们的Wnt信号传导活性。我们证明，LRP 6C具有很强的信号传导活性，而LRP 5C在细胞中的活性要低得多。重组LRP 5C和LRP 6C在体外磷酸化后表现出相似的Axin结合能力，表明LRP 5和LRP 6在体内在Axin结合之前的步骤（可能在接受磷酸化时）不同。我们在两个最羧基的PPPSPxS基序之间鉴定了一个插入的“gap 4”区域，该区域似乎解释了LRP 5C和LRP 6C之间的大部分差异，并表明该区域的改变足以增强LRP 5 PPPSPxS磷酸化和信号传导至与细胞中的LRP 6相当的水平。此外，我们提供的证据表明磷酸化LRP 5或LRP 6与Axin的结合可能是直接的，不需要GSK 3激酶作为桥接中间体。因此，我们的研究揭示了一个新的和重要的分子调节机制的差异调节LRP 5和LRP 6的磷酸化和信号转导活性。
Low-density lipoprotein receptor-related proteins 5 and 6 (LRP5 and LRP6) serve as Wnt co-receptors for the canonical β-catenin pathway. While LRP6 is essential for embryogenesis, both LRP5 and LRP6 play critical roles for skeletal remodeling, osteoporosis pathogenesis and cancer formation, making LRP5 and LRP6 key therapeutic targets for cancer and disease treatment. LRP5 and LRP6 each contain in the cytoplasmic domain five conserved PPPSPxS motifs that are pivotal for signaling and serve collectively as phosphorylation-dependent docking sites for the scaffolding protein Axin. However existing data suggest that LRP6 is more effective than LRP5 in transducing the Wnt signal. To understand the molecular basis that accounts for the different signaling activity of LRP5 and LRP6, we generated a series of chimeric receptors via swapping LRP5 and LRP6 cytoplasmic domains, LRP5C and LRP6C, and studied their Wnt signaling activity using biochemical and functional assays. We demonstrate that LRP6C exhibits strong signaling activity while LRP5C is much less active in cells. Recombinant LRP5C and LRP6C upon in vitro phosphorylation exhibit similar Axin-binding capability, suggesting that LRP5 and LRP6 differ in vivo at a step prior to Axin-binding, likely at receiving phosphorylation. We identified between the two most carboxyl PPPSPxS motifs an intervening “gap4” region that appears to account for much of the difference between LRP5C and LRP6C, and showed that alterations in this region are sufficient to enhance LRP5 PPPSPxS phosphorylation and signaling to levels comparable to LRP6 in cells. In addition we provide evidence that binding of phosphorylated LRP5 or LRP6 to Axin is likely direct and does not require the GSK3 kinase as a bridging intermediate as has been proposed. Our studies therefore uncover a new and important molecular tuning mechanism for differential regulation of LRP5 and LRP6 phosphorylation and signaling activity.
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