LRP6 phosphorylation in Wnt/beta-catenin signaling

描述（由申请人提供）：通过经典α-连环蛋白途径的Wnt信号传导控制细胞命运决定和细胞增殖，并且对动物发育至关重要。Wnt/β-连环蛋白信号传导缺陷与人类结直肠癌、家族性骨质疏松症和其他疾病有关。因此，了解Wnt/b-连环蛋白信号传导与人类健康高度相关。Wnt/β-连环蛋白信号传导由2个不同的细胞表面受体家族启动。一种（1）是蛇形受体的卷曲（Fz）家族的成员，另一种是LDL受体相关蛋白家族的单跨膜受体，LRP 5或LRP 6。Wnt如何导致这些受体的激活是一个关键但知之甚少的问题。何博士的研究小组表明，Fz和LRP 5/6在体外形成了Wnt诱导的辅助受体复合物，LRP 6在信号传导过程中起着关键作用。他们最近发现磷酸化可能是LRP 6激活的基础。他们发现PPP（S/T）P基序在LRP 5/6的细胞内结构域中重复5次，对LRP 6信号传导功能至关重要。他们证明，PPP（S/T）P基序的磷酸化是信号传导和Axin（一种调节b-连环蛋白稳定性的关键支架蛋白）结合/识别所必需的。他们进一步表明，Wnt在体内诱导LRP 6磷酸化。这些结果表明，Wnt通过诱导LRP 6磷酸化激活跨膜信号传导。在这个应用程序中，3个具体的目标是为了充实/扩展这个工作模型。(1)全面表征LRP 6胞内结构域的所有五个PPP（S/T）P基序。待解决的问题包括这些PPP（S/T）P基序的功能，以及它们的差异磷酸化是否控制LRP 6信号。(2)研究磷酸化PPP（S/T）P基序与Axin的相互作用。目的是鉴定识别PPP（S/T）P磷酸化的Axin结构域/模块，并检查LRP 6-Axin缔合是否改变Axin复合物的组成，从而控制b-连环蛋白稳定性。(3)鉴定磷酸化PPP（S/T）P基序的激酶。目的是研究已知的Wnt/α-连环蛋白信号转导相关激酶在PPP（S/T）P磷酸化中的潜在作用，并通过几种互补方法分离PPP（S/T）P激酶或激酶。这些实验将显著增强对发育和疾病中Wnt受体激活和Wnt信号转导的理解。

DESCRIPTION (provided by applicant): Wnt signaling through the canonical a-catenin pathway controls cell fate determination and cell proliferation, and is essential for animal development. Defective Wnt/b-catenin signaling has been associated with human colorectal cancer, familial osteoporosis, and other diseases. Thus understanding Wnt/b-catenin signaling is highly relevant to human health. Wnt/b-catenin signaling is initiated by 2 distinct families of cell surface receptors. One (1) is a member of the Frizzled (Fz) family of serpentine receptors, and the other is a single transmembrane receptor of the LDL receptor related protein family, LRP5 or LRP6. How Wnt leads to the activation of these receptors is a critical but poorly understood issue. Dr. He's group showed that Fz and LRP5/6 form a Wnt-inducible co-receptor complex in vitro, and that LRP6 plays a key role in the signaling process. They recently discovered that phosphorylation likely underlies LRP6 activation. They found that a PPP(S/T)P motif, which is reiterated 5 times in the intracellular domain of LRP5/6, is essential for LRP6 signaling function. They demonstrated that phosphorylation of this PPP(S/T)P motif is required for signaling and for binding/recognition by Axin, a key scaffolding protein that regulates b-catenin stability. They further showed that Wnt induces LRP6 phosphorylation in vivo. These results suggest that Wnt activates transmembrane signaling via inducing LRP6 phosphorylation. In this application, 3 specific aims are designed to substantiate/extend this working model. (1) To characterize fully all five PPP(S/T)P motifs in LRP6 intracellular domain. Issues to be addressed include the function of these PPP(S/T)P motifs and whether their differential phosphorylation controls LRP6 signaling. (2) To investigate Axin interaction with the phosphorylated PPP(S/T)P motifs. The aim is to identify the Axin domain/module that recognizes PPP(S/T)P phosphorylation, and examine whether LRP6-Axin association alters the composition of the Axin complex, thereby governing b-catenin stability. (3) To identify kinase (or kinases) that phosphorylates the PPP(S/T)P motif. This aim is to investigate known kinases implicated in Wnt/a-catenin signaling for their potential roles in PPP(S/T)P phosphorylation and to isolate the PPP(S/T)P kinase or kinases via several complementary approaches. These experiments will significantly enhance the understanding of Wnt receptor activation and Wnt signal transduction in development and disease.