The Role of Merlin Phosphorylation on its Tumor Suppressive Activity

描述（由申请人提供）：NF 2肿瘤抑制基因编码一种称为merlin或schwannomin的细胞内膜相关蛋白，属于将细胞表面糖蛋白与肌动蛋白细胞骨架连接的细胞骨架相关蛋白的带4.1家族（ERM蛋白）。Merlin生长抑制依赖于其形成有效N端/C端结合的能力。Merlin以“开放”（非活性形式）和“封闭”（活性生长抑制形式）构象存在，这些构象受到蛋白质N末端和C末端之间分子内结合的调节。大量研究表明，梅林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merlin，而不是患者衍生的突变体（L 64 P），选择性地结合PIKE-L和抑制磷酸肌醇3-激酶（PI 3-激酶）的活性。PIKE（PI 3-Kinase Enhancer）是一种脑特异性的GT3，其结合PI 3-kinase并刺激其脂质激酶活性。PI 3-激酶活性的这种抑制是由于merlin破坏PIKE-L与PI 3-激酶的结合。merlin的诱导显著减少了神经鞘瘤细胞中PI 3-激酶/Akt信号通路。磷酸化和磷酸肌醇脂质结合协调介导ERM蛋白的活化。然而，Akt是否磷酸化merlin和磷酸肌醇脂质结合merlin也控制merlin活性仍然难以捉摸。我们推测merlin是通过PI 3-激酶/Akt级联负反馈机制进行生理调节的。具体来说，我们将测试Akt是否磷酸化梅林和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，从而消除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理解merlin的生理功能至关重要，而且对于未来开发治疗这种疾病的新药也至关重要。

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.