Filamin interactions in differentiation, invasion and disease

描述（由申请人提供）：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。丝状体是细胞中 由N-末端肌动蛋白结合结构域和24个免疫球蛋白样结构域组成的交联蛋白，其与许多胞质信号蛋白和跨膜受体相互作用。人类有三种细丝蛋白基因，编码广泛表达的细丝蛋白A和B以及主要是肌肉特异性细丝蛋白C。在film中的错义点突变引起多种人类疾病，范围从改变的神经元迁移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、心脏缺陷和阻塞性尿路病。肌动蛋白结合域是细丝蛋白突变的热点，但是，尽管在理解细丝蛋白结构和功能方面取得了巨大进展，细丝蛋白点突变如何引起疾病仍然知之甚少。此外，细丝蛋白A表达减少与乳腺癌侵袭和转移增加相关，我们最近发现细丝蛋白的丢失增加细胞外基质（ECM）重塑和细胞侵袭。细丝蛋白如何控制ECM降解和侵袭尚不清楚。此外，我们已经表明，ASB 2（锚蛋白重复含有蛋白与细胞因子信号传导盒2的抑制剂），E3泛素连接酶复合物的一部分，目标filaminin快速蛋白酶体降解，我们认为，由此产生的瞬时损失的细丝蛋白有助于维甲酸诱导的急性早幼粒细胞白血病细胞的分化。然而，ASB 2功能的分子基础以及细丝蛋白的缺失如何影响细胞分化尚未阐明。为了解决上述重要的未回答的问题，我们提出了三个具体的目标，利用我们丰富的经验，利用生物化学，细胞和结构技术来调查filmosphere。具体而言，我们将：1）表征ASB 2介导的细丝蛋白降解的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;和3）鉴定与疾病相关的细丝蛋白点突变相关的细胞表型，揭示疾病的潜在分子机制。

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.