The Role of Merlin Phosphorylation on its Tumor Suppressive Activity

描述(申请人提供)：NF2肿瘤抑制基因编码一种细胞内膜相关蛋白，称为Merlin或Schwannomin，属于4.1带(ERM蛋白)细胞骨架相关蛋白家族，将细胞表面糖蛋白与肌动蛋白细胞骨架联系起来。梅林生长抑制取决于其形成生产性N-Term/C-Term关联的能力。Merlin以“开放”(非活性形式)和“封闭”(活性生长抑制形式)构象存在，由蛋白质的N-末端和C-末端之间的分子内结合来调节。大量研究表明，Merlin磷酸化在决定其构象和结合活性方面起着至关重要的作用。最近，我们发现野生型的Merlin，而不是患者来源的突变体(L64P)，选择性地与Pike-L结合，并抑制磷酸肌醇3-激酶(PI-3-Kinase)的活性。Pike(PI 3-Kinase Enhancer)是一种脑特异性GTP酶，能与PI 3-激酶结合并刺激其脂激酶活性。这种对PI3K活性的抑制是由于Merlin破坏了Pike-L与PI3K的结合。在神经鞘瘤细胞中，Merlin的诱导显著降低了PI3-K/Akt信号通路。磷酸化和肌醇磷脂结合协同介导ERM蛋白的激活。然而，Akt是否使Merlin磷酸化以及与Merlin结合的肌醇磷脂是否也控制Merlin的活性仍不清楚。我们推测，Merlin在生理上受PI3-K/Akt级联负反馈机制的调节。具体地说，我们将测试Akt是否使Merlin磷酸化，并调节其构象和亚细胞定位。此外，我们将确定Akt磷酸化和PI(3，4和5)P3是否协同介导Merlin与关键下游效应物的结合活性，从而取消Merlin的肿瘤抑制活性。鉴定介导Merlin磷酸化和结合活性的信号通路不仅对于了解Merlin的生理功能，而且对于未来开发治疗该病的新药也是至关重要的。

DESCRIPTION (provided by applicant): The NF2 tumor suppressor gene encodes an intracellular membrane-associated protein, called merlin or schwannomin, which belongs to the band 4.1 family (ERM proteins) of cytoskeleton-associated proteins that link cell surface glycoproteins to the actin cytoskeleton. Merlin growth suppression is dependent on its ability to form a productive N-term/C-term association. Merlin exists in "open" (inactive form) and "closed" (active growth suppressive form) conformations, which are regulated by an intramolecular association between the N- and C-termini of the protein. Numerous studies demonstrate that merlin phosphorylation plays an essential role in dictating its conformation and binding activity. Recently, we showed that wild-type merlin, but not patient-derived mutant (L64P), selectively binds PIKE-L and inhibits Phosphoinositol 3-kinase (PI 3- kinase) activity. PIKE (PI 3-Kinase Enhancer) is a brain-specific GTPase that binds to PI 3-kinase and stimulates its lipid kinase activity. This suppression of PI 3-kinase activity results from merlin disrupting the binding of PIKE-L to PI 3-kinase. Induction of merlin substantially diminishes PI 3-kinase/Akt signaling pathway in Schwannoma cells. Phosphorylation and phosphoinositol lipids binding coordinately mediate the activation of ERM proteins. However, whether Akt phosphorylates merlin and phosphoinositol lipids bind to merlin also control merlin activity remains elusive. We hypothesize that merlin is physiologically regulated by a negative feed-back mechanism of PI 3-kinase/Akt cascade. Specifically, we will test whether Akt phosphorylates merlin and modulates its conformation and subcellular localization. In addition, we will determine whether Akt phosphorylation and PI (3, 4, and 5) P3 synergistically mediate merlin's binding activity to the critical downstream effectors, resulting in abolishing merlin's tumor suppressive activity. Identification of signaling pathways mediating merlin phosphorylation and binding activity is essential not only for understanding the physiological functions of merlin, but also for the future development of novel drug treatments for this disease.