Abstract：
Fascin, a major actin cross-linking protein, is expressed in most vertebrate epithelial tissues. It organizes actin filaments into well-ordered bundles that are responsible for the extension of dynamic membrane protrusions, including microspikes, filopodia, and invadopodia from cell surfaces, which are involved in cell migration and invasion as critical components of cancer metastasis. However, it is not well-understood how fascin-1 induces actin binding/bundling and where fascin-1 localizes along the actin filaments, thus facilitating actin bundle formation. In the present study, we attempted to clarify these problems by using biochemical and electron microscopic analyses using various fascin-1 constructs. Three dimensional structures of actin/fascin-1 complex were obtained by electron microscopy (EM) with iterative helical real-space reconstruction (IHRSR) and tomography. We revealed that the N-terminal region containing the Actin-Binding Site 2 (ABS2) of fascin-1 is responsible for actin bundling and the C-terminal region is important for the dimerization of fascin-1. We also found that the dimerization of fascin-1 through intermolecular interactions of the C-terminal region is essential for actin bundling. Since fascin is an important factor in cancer development, it is expected that the findings of present study will provide useful information for development of therapeutic strategies for cancer.
摘要：
Fascin,一种主要的肌动蛋白交联蛋白，在大多数脊椎动物上皮组织中表达。它将肌动蛋白丝组织成有序的束，这些束负责动态膜突起的延伸，包括微尖峰,filopodia,和从细胞表面侵入足印，它们参与细胞迁移和侵袭，是癌症转移的关键组成部分。然而,尚不清楚fascin-1如何诱导肌动蛋白结合/成束，以及fascin-1沿肌动蛋白丝定位的位置，从而促进肌动蛋白束的形成。在本研究中,我们试图通过使用各种fascin-1构建体的生化和电子显微镜分析来阐明这些问题。通过电子显微镜（EM），迭代螺旋实空间重建（IHRSR）和断层扫描获得了肌动蛋白/fascin-1复合物的三维结构。我们发现，包含fascin-1的肌动蛋白结合位点2（ABS2）的N末端区域负责肌动蛋白捆扎，而C末端区域对于fascin-1的二聚化很重要。我们还发现，通过C末端区域的分子间相互作用使fascin-1二聚化对于肌动蛋白捆扎至关重要。由于fascin是癌症发展的重要因素，预计本研究的结果将为癌症治疗策略的制定提供有用的信息.
