EFFECT OF PHOSPHORYLATION ON INTERACTION OF P130CAS WITH AND-34

磷酸化对 P130CAS 与 AND-34 相互作用的影响

• 批准号: 8365552
• 负责人: ADAM LERNER
• 金额: $ 1.54万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-06-01 至 2012-08-09
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8365552
• 关键词: Adhesions; Amino Acids; BCAR3 gene; Biology; Breast Cancer Cell; Cancer Cell Growth; Cancer cell line; Cells; Employee Strikes; Epithelial; Estrogen Antagonists; Family member; Fibronectins; Focal Adhesions; Funding; Grant; Growth; Hour; MCF7 cell; Mass Spectrum Analysis; Mediating; Medicine; Mesenchymal; National Center for Research Resources; Pattern; Phenotype; Phosphorylation; Post-Translational Protein Processing; Principal Investigator; Proline-Rich Domain; Protein Family; Proteins; Publishing; Research; Research Infrastructure; Resistance; Resources; SH2D3A gene; SH2D3C gene; Serine; Signal Transduction; Sodium Dodecyl Sulfate-PAGE; Source; Tyrosine Phosphorylation; United States National Institutes of Health; adapter protein; cost; human BCAR1 protein; small hairpin RNA; src-Family Kinases

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. This project seeks to establish why, upon over-expression of a specific protein, AND-34, NSP protein family members associate with p130Cas, a focal adhesion adapter protein best known as a Src substrate that integrates adhesion-related signaling. Over-expression of AND-34/BCAR3/NSP2 (BCAR3), but not NSP1 or NSP3, induces anti-estrogen resistance in human breast cancer cell lines. BCAR3 over-expression in epithelial MCF-7 cells augments levels of a phosphorylated p130Cas species that migrates more slowly on SDS PAGE while NSP-1 and NSP3 induce modest or no phosphorylation, respectively. Conversely, reduction in BCAR3 expression in mesenchymal MDA-231 cells by inducible shRNA results in loss of such p130Cas phosphorylation. Replacement of NSP3's serine/proline-rich domain with that of AND- 34/BCAR3 instills the ability to induce p130Cas phosphorylation. Phospho-amino acid analysis demonstrates that BCAR3 induces p130Cas serine phosphorylation. Mass spectrometry identified phosphorylation at p130Cas serines 139, 437 and 639. p130Cas serine phosphorylation occurs physiologically hours after adhesion of MDA-231 cells to fibronectin and is dependent upon BCAR3 expression. BCAR3 knockdown also alters p130Cas localization and converts MDA-231 growth to an epithelioid pattern characterized by striking cohesiveness and lack of lamellipodial projections at colony borders. These studies suggest that BCAR3 regulates p130Cas serine phosphorylation that is adhesion-dependent, temporally distinct from previously well-characterized rapid Fak and Src kinase-mediated p130Cas tyrosine phosphorylation and that correlates with invasive phenotype. These results were published in Cellular Signalling (Makkinje A, Near RI, Infusini G, Vanden Borre P, Bloom A, Cai D, CostelloCE, Lerner A. AND-34/BCAR3 regulates adhesion-dependent p130Cas serine phosphorylation andbreast cancer cell growth pattern.Cell Signal. A Makkinje et al., 2009, 21, 1423-1435). Current studies aim at identifying protein partners and their post-translational modifications.

该副本是利用资源的众多研究子项目之一 由NIH/NCRR资助的中心赠款提供。对该子弹的主要支持 而且，副投影的主要研究员可能是其他来源提供的 包括其他NIH来源。 列出的总费用可能 代表subproject使用的中心基础架构的估计量， NCRR赠款不直接向子弹或副本人员提供的直接资金。 该项目旨在确定为什么在特定蛋白质过度表达的情况下，以及-34，NSP蛋白质家族成员与P130CAS相关，这是一种局灶性粘附衔接子蛋白，最著名的是整合与粘附相关的信号传导的SRC底物。 And-34/BCAR3/NSP2（BCAR3）的过表达，而不是NSP1或NSP3，可诱导人类乳腺癌细胞系中的抗雌激素耐药性。上皮MCF-7细胞中的BCAR3过表达增加了磷酸化的P130CAS物种的水平，该物种在SDS页面上迁移较慢，而NSP-1和NSP3分别诱导适度或没有磷酸化。相反，可诱导的shRNA在间充质MDA-231细胞中降低BCAR3表达导致这种P130CAS的损失 磷酸化。用和 - 替换NSP3的丝氨酸/脯氨酸富含域 34/BCAR3灌输了诱导P130CAS磷酸化的能力。磷酸氨基酸分析表明，BCAR3诱导P130CAS丝氨酸磷酸化。质谱法在P130CAS丝氨酸139、437和639中鉴定出磷酸化。P130CAS丝氨酸磷酸化在MDA-231细胞粘附后在生理上发生的磷酸化发生在生理上，并依赖于BCAR3表达。 BCAR3敲低还改变了P130CAS的定位，并将MDA-231的增长转化为上皮细胞的模式，其特征是具有醒目的凝聚力和缺乏薄膜 殖民地边界的预测。这些研究表明，BCAR3调节粘附依赖性的P130CAS丝氨酸磷酸化，在时间上与以前特征良好的快速FAK和SRC激酶介导的P130CAS酪氨酸磷酸化，并且与侵入性表型相关。这些结果发表在细胞信号传导中（Makkinje A，RI附近，Infusini G，Vanden Borre P，Bloom A，Cai D，Costelloce，LernerA。当前的研究旨在鉴定蛋白质伴侣及其翻译后修饰。