Elucidating the role of palladin/alpha-actinin in cell motility and invasion

描述(申请人提供)：癌症的特征是细胞具有侵袭性、侵袭性，并且通常是转移的。只有一小部分肿瘤细胞具有转移的能力，然而正是转移的亚群使癌症成为一种致命的疾病。因此，了解这类细胞与正常肿瘤细胞有何不同是至关重要的。如果我们能找到转移性细胞的特异性标记，那么我们就有希望杀死它们，或者至少限制它们。这项建议将集中在Palladin和a-Actinin的确切分子功能上，这两种蛋白质都被认为在细胞运动中发挥关键作用，特别是在癌症转移中。新的蛋白Palladin是一种多结构域的肌动蛋白相关蛋白，在脊椎动物中高度保守。Palladin基因敲除小鼠表现出胚胎致死的表型，证实了Palladin在哺乳动物胚胎发生中起着至关重要的作用，Palladin缺失细胞在运动性和粘附性方面都存在缺陷。α-肌动蛋白还参与肌动蛋白的交联，是细胞信号转导和肌动蛋白重组所涉及的多组分复合体组装的平台。最近，人类Palladin的一个突变(P239S突变)被认为与一种遗传性胰腺癌有关，这表明Palladin功能受损可能与一种高度侵袭性的人类疾病有关。这种突变也发生在与α-肌动蛋白结合所需的palladin区域。因此，Palladin和α-Actinin在功能和物理上都与正常和病理细胞的运动有关。目前的研究集中在使用Palladin或α-Actinin水平作为预后生物标记物。然而，我们的假设是，Palladin/a-Actinin复合体是转移性癌症的相关生物标记物，并希望这项工作将导致最致命的癌症的预后标记物得到改善。目前，该复合体在组织肌动蛋白细胞骨架中的确切分子作用尚不清楚。我们建议用核磁共振来解决溶液中的复合体的结构，这反过来将指导我们对Palladin的合理突变来破坏复合体。然后，我们将利用对培养的乳腺癌细胞的转染来表征缺乏结合α-肌动蛋白能力的palladin突变体的生物学活性。总之，这些目标将使我们能够确定Palladin和α-Actinin之间的相互作用在细胞运动和侵袭中的生物学意义。

DESCRIPTION (provided by applicant): Cancer is characterized by cells that are aggressive, invasive, and often metastatic. Only a subset of tumor cells possesses the ability to metastasize, and yet it is the metastatic sub-population that makes cancer a lethal disease. Therefore, it is critical to understand how this population of cells differs from normal tumor cells. If we can find markers that are specific to the metastatic cells, then there is a hope that we can kill them, or at least restrict them. This proposal will focus on the precise molecular function of palladin and a-actinin, two proteins that have both been implicated as playing critical roles in cell motility and particularly in cancer metastasis. The novel protein palladin is a multi-domain, actin-associated protein that is highly conserved among vertebrate species. The palladin knockout mouse displays a phenotype of embryonic lethality, confirming that palladin plays an essential role in mammalian embryogenesis, and palladin null cells have defects in both motility and adhesion. a-Actinin is also involved in actin-crosslinking and serves as a platform for the assembly of multi-component complexes involved in cell signaling and actin reorganization. Recently, a mutation in human palladin (the P239S mutation) was implicated in an inherited form of pancreatic cancer, suggesting that impaired palladin function may contribute to a highly invasive human disease. This mutation also occurs in a region of palladin required for binding to a-actinin. Therefore palladin and a-actinin are both functionally and physically linked to both normal and pathological cell motility. Current research is focused on using palladin or a-actinin levels as prognostic biomarkers. Yet, our hypothesis is that the palladin/a-actinin complex is the relevant biomarker for metastatic cancer and hope that this work will lead to improved prognostic markers for the deadliest of cancers. Currently the precise molecular role of this complex in organizing the actin cytoskeleton is unknown. We propose to solve the structure of the complex in solution using nuclear magnetic resonance, which will in turn guide our rational mutagenesis of palladin to disrupt the complex. Then we will use transfection of cultured breast cancer cells to characterize the biological activity of palladin mutants that are deficient in their ability to bind a-actinin. Together, these aims will allow us to determine the biological significance of the interaction between palladin and a-actinin in cell motility and invasion.