Molecular Mechanisms Of Cell Adhesion And Invasion

细胞粘附和侵袭的分子机制

• 批准号: 7967969
• 负责人: Steven Akiyama
• 金额: $ 101.92万
• 依托单位: NATIONAL INSTITUTE OF ENVIRONMENTAL HEALTH SCIENCES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7967969
• 关键词: 1-Phosphatidylinositol 3-Kinase; Acute; Acute Disease; Adhesions; Adhesives; Affinity Chromatography; Animals; Autoimmunity; Automobile Driving; Basement membrane; Binding; Biological Models; Blood Platelets; Blood Vessels; Bone remodeling; CD29 Antigen; Cell Adhesion; Cell Adhesion Molecules; Cell Line; Cell Surface Receptors; Cell membrane; Cell surface; Cell-Cell Adhesion; Cells; Chimeric Proteins; Chronic; Collagen; Complex; Cues; Cytoskeleton; Data; Development; Disease; Disease Progression; E-Selectin; Embryo; Embryonic Development; Endogenous Factors; Endothelial Cells; Endothelium; Event; Exposure to; Extracellular Matrix; Family; Fibronectins; Gelatinase B; Glycoproteins; Goals; Human; Immunoglobulin G; Inflammation; Inflammatory; Inflammatory Response; Integrin-mediated Cell Adhesion Pathway; Integrins; Intervention; L-Selectin; Laminin; Leukocyte Rolling; Leukocyte Trafficking; Leukocytes; Life; MAP Kinase Gene; MAPK14 gene; Malignant Neoplasms; Mediating; Melanoma Cell; Membrane Glycoproteins; Membrane Proteins; Molecular; Morphogenesis; Neoplasm Metastasis; P-Selectin; Pathway interactions; Phenotype; Play; Post-Translational Protein Processing; Prevention strategy; Process; Proteins; Proteomics; Recombinants; Regulation; Research; Role; Selectins; Side; Signal Pathway; Signal Transduction; Signal Transduction Pathway; Site; Small Interfering RNA; Stimulus; System; Thrombin; Thrombosis; Tissues; Toxic Environmental Substances; Tyrosine Phosphorylation; Wound Healing; adhesion receptor; cell motility; crosslink; design; exposed human population; extracellular; human disease; human migration; implantation; member; migration; mitogen-activated protein kinase p38; neoplastic cell; novel; nucleolin; receptor; response

Cell adhesion and migration contribute to normal processes such as differentiation, embryonic development, and wound healing as well as to the progression of diseases and pathological conditions that can result from either acute or chronic exposure to environmental toxicants, such as cancer and inflammatory responses. Key mechanistic steps in these processes involve the interactions of extracellular glycoproteins--such as fibronectin, laminin, and collagens--with specific adhesive receptors, the best characterized of which are the integrins, a family of heterodimeric complexes consisting of an alpha subunit and a beta subunit. Integrins are highly regulated receptors that can exist in either an active or inactive state. Selectins are vascular cell-cell adhesion molecules involved in leukocyte trafficking, inflammation, thrombosis, autoimmunity and cancer. Accumulation of leukocytes at sites of inflammation is initiated by selectins that mediate the capturing and rolling of leukocytes on endothelium. Three major members of the selectin family have been identified: L-selectin, E-selectin and P-selectin. L-Selectin is constitutively expressed on leukocytes. P-and E-selectins are expressed on activated endothelial cells in response to microenvirnomental stimuli. P-selectin is also expressed on thrombin-activated platelets. All three members of the selectin family, E-, L-, and P-selectin can bind to human tumor cells and cancer-derived cell line. Our research has focused recently on the possible role of inflammatory cues in activation of integrin-mediated tumor cell migration and invasion. As a model system, we are examining the ability of one selectin, P-selectin, to trigger integrin-mediated adhesion and migration of cultured human tumor cells. We focus on two closely related aspects of this project: to characterize the mechanisms of P-selectin-induced activation of integrin-mediated cell adhesion and cell migration. We have previously shown that binding of soluble, recombinant P-selectin-IgG Fc chimeric protein to Colo-320 cells stimulates cell adhesion to fibronectin through the specific activation of the alpha5-beta1 integrin by means of the p38 MAP kinase and PI-3 kinase (PI3-k) signaling pathways. We have now identified nucleolin as a novel cell surface P-selectin receptor on Colo-320 cells using affinity chromatography and a proteomic approach. This finding was validated by showing that an anti-nucleolin mAb D3 inhibits P-selectin interactions with living Colo-320 cells and that nucleolin becomes clustered at the external side of the plasma membrane of living, intact cells when bound to cross-linked P-selectin-IgG-Fc chimeric protein. We have also found that P-selectin binding to Colo-320 cells induces tyrosine phosphorylation specifically of cell-surface nucleolin and formation of a signaling complex containing cell surface nucleolin, PI 3-K, and p38 MAPK. Using siRNA approaches, we showed that both P-selectin binding to Colo-320 cells and formation of the P-selectin-mediated p38 MAPK/ PI 3-K signaling complex require nucleolin expression. Thus, we have characterized nucleolin (or a nucleolin-like protein) as a novel, signaling cell-surface receptor for P-selectin on Colo-320 cells and suggest a mechanism for linkage of nucleolin to P-selectin-induced signal transduction pathways that regulate the adhesion of Colo-320 on fibronectin substrates. We have recently been characterizing the stimulation of integrin-mediated tumor cell migration by the binding of soluble, recombinant P-selectin-IgG Fc chimeric protein. We have found that P-selectin stimulates the secretion of matrix metalloproteinase-9 (MMP-9) by A375 human melanoma cells and that direct addition of exogenous MMP-9 to A-375 cells can stimulate the migratory phenotype. The current paradigm in the field states that MMP-9 stimulates migration by digesting basement membrane proteins, allowing tumor cells to clear a migratory pathway. However, we have preliminary data that show that stimulation of cell migration by MMP-9 can occur even in the absence of catalytic activity.

细胞粘附和迁移有助于正常过程，如分化、胚胎发育和伤口愈合，以及可能由急性或慢性暴露于环境毒物引起的疾病和病理状况的进展，如癌症和炎症反应。这些过程中的关键机制步骤涉及细胞外糖蛋白-如纤连蛋白，层粘连蛋白和胶原蛋白-与特异性粘附受体的相互作用，其中最具特征的是整合素，一个由α亚基和β亚基组成的异二聚体复合物家族。整合素是高度调节的受体，可以以活性或非活性状态存在。 选择素是血管细胞-细胞粘附分子，参与白细胞运输、炎症、血栓形成、自身免疫和癌症。白细胞在炎症部位的积聚由介导白细胞在内皮上的捕获和滚动的选择素引发。 选择素家族的三个主要成员已被鉴定：L-选择素、E-选择素和P-选择素。L-选择素在白细胞上组成型表达。P-选择素和E-选择素在微环境刺激下在活化的内皮细胞上表达。P-选择素也在凝血酶活化的血小板上表达。 选择素家族的所有三个成员，E-、L-和P-选择素可以与人肿瘤细胞和癌源性细胞系结合。 我们的研究最近集中在炎症因子在激活整合素介导的肿瘤细胞迁移和侵袭中的可能作用。作为一个模型系统，我们正在研究一种选择素，P-选择素，触发整合素介导的粘附和迁移的培养的人肿瘤细胞的能力。我们专注于这个项目的两个密切相关的方面：P-选择素诱导激活整合素介导的细胞粘附和细胞迁移的机制的特点。 我们先前已经证明，可溶性重组P-选择素-IgG Fc嵌合蛋白与科洛-320细胞的结合通过p38 MAP激酶和PI-3激酶（PI 3-k）信号传导途径特异性激活α 5-β 1整联蛋白来刺激细胞与纤连蛋白的粘附。 我们现在已经确定核仁素作为一种新的细胞表面P-选择素受体科洛-320细胞使用亲和层析和蛋白质组学方法。通过显示抗核仁素mAb D3抑制P-选择素与活的科洛-320细胞的相互作用，并且当与交联的P-选择素-IgG-Fc嵌合蛋白结合时，核仁素在活的完整细胞的质膜外侧聚集，验证了该发现。 我们还发现P-选择素与科洛-320细胞的结合诱导细胞表面核仁素的酪氨酸磷酸化，并形成含有细胞表面核仁素、PI 3-K和p38 MAPK的信号复合物。 利用siRNA方法，我们发现P-选择素与科洛-320细胞的结合以及P-选择素介导的p38 MAPK/ PI 3-K信号复合物的形成都需要核仁素的表达。因此，我们的特点是核仁素（或核仁素样蛋白）作为一种新的，信号细胞表面受体的P-选择素对科洛-320细胞，并提出了一种机制，连接核仁素的P-选择素诱导的信号转导途径，调节粘附的科洛-320纤连蛋白基板。 我们最近已经表征了通过可溶性重组P-选择素-IgG Fc嵌合蛋白的结合来刺激整合素介导的肿瘤细胞迁移。 我们已经发现P-选择素刺激A375人黑素瘤细胞分泌基质金属蛋白酶-9（MMP-9），并且直接向A-375细胞添加外源性MMP-9可以刺激迁移表型。 目前该领域的研究范式表明，MMP-9通过消化基底膜蛋白刺激迁移，使肿瘤细胞清除迁移途径。 然而，我们有初步的数据表明，刺激细胞迁移的MMP-9可以发生，即使在催化活性的情况下。