Filamin interactions in differentiation, invasion and disease

描述(由申请人提供)：我们要求继续支持我们对细丝蛋白在细胞分化、侵袭和疾病中的作用的研究。丝蛋白是必不可少的肌动蛋白 交联蛋白由一个N-末端肌动蛋白结合区和24个免疫球蛋白样区组成，它们与许多胞浆信号蛋白和跨膜受体相互作用。人类有三个细丝蛋白基因，编码广泛表达的细丝蛋白A和B，主要是肌肉特有的细丝蛋白C。丝蛋白错义点突变会导致多种人类疾病，从神经元迁移改变到心脏和骨骼肌缺陷，以及一系列先天性畸形，通常以骨骼发育不良为特征，但也包括骨骼外畸形，如腭裂、心脏缺陷和梗阻性尿病。肌动蛋白结合区是细丝蛋白突变的研究热点，但是，尽管在了解细丝蛋白的结构和功能方面取得了巨大的进展，但细丝蛋白点突变如何导致疾病仍然知之甚少。此外，细丝蛋白A的表达减少与乳腺癌侵袭和转移增加有关，我们最近发现，细丝蛋白的缺失会增加细胞外基质(ECM)的重塑和细胞侵袭。丝素如何控制细胞外基质的降解和侵袭尚不清楚。此外，我们还发现，作为E3泛素连接酶复合体的一部分，ASB2(Ankyrin Repeat-Containing Protein with a Inhibitor of cell Kine Signal Box2)是E3泛素连接酶复合体的一部分，它以丝蛋白为靶标快速降解蛋白酶体，我们认为，由此导致的丝蛋白的瞬时丢失有助于维甲酸诱导急性早幼粒细胞白血病细胞的分化。然而，ASB2功能的分子基础以及细丝蛋白的丢失如何影响细胞分化还没有详细阐述。为了解决上面强调的重要的悬而未决的问题，我们提出了三个具体的目标，利用我们使用生化、细胞和结构技术来研究丝蛋白的丰富经验。具体地说，我们将：1)表征ASB2介导的细丝降解机制并测试其在细胞分化中的作用；2)评估细丝蛋白在EC重塑和细胞侵袭中的作用；以及3)确定与疾病相关的细丝蛋白点突变相关的细胞表型，揭示潜在的疾病分子机制。

DESCRIPTION (provided by applicant): Continued support is requested for our investigation of the roles of filamin in cell differentiation, invasion and disease. Filamins are essential actin crosslinking proteins composed of an N-terminal actin-binding domain followed by 24 immunoglobulin-like domains which interact with numerous cytosolic signaling proteins and transmembrane receptors. Humans have three filamin genes, encoding the widely expressed filamin A and B and largely muscle specific filamin C. Missense point mutations in filamins cause a variety of human diseases, ranging from altered neuronal migration, to cardiac and skeletal muscle defects, and a spectrum of congenital malformations generally characterized by skeletal dysplasias but also including extra-skeletal malformations such as cleft palate, cardiac defects and obstructive uropathy. The actin-binding domain is a hotspot for filamin mutations but, despite dramatic progress in understanding filamin structure and function, how filamin point mutations cause disease remains poorly understood. Furthermore, reduced filamin A expression correlates with increased breast cancer invasion and metastasis, and we recently discovered that loss of filamin increases extracellular matrix (ECM) remodeling and cell invasion. How filamin controls ECM degradation and invasion is unknown. In addition, we have shown that ASB2 (ankyrin repeat-containing protein with a suppressor of cytokine signaling box 2), part of an E3 ubiquitin ligase complex, targets filamins for rapid proteasomal degradation and we suggest that the resultant transient loss of filamin contributes to retinoic acid-induced differentiation of acute promyelocytic leukemia cells. However, the molecular basis for ASB2 function and how loss of filamin influences cell differentiation have not been elaborated. To address the important unanswered questions highlighted above we propose three specific aims which draw on our extensive experience using biochemical, cellular and structural techniques to investigate filamins. Specifically, we will: 1) Characterize the mechanism of ASB2-mediated filamin-degradation and test its role in cell differentiation; 2) Assess the role of filamins in EC remodeling and cell invasion; and 3) Identify cellular phenotypes associated with disease-associated filamin point mutations, revealing potential molecular mechanisms of disease.