Role of CEACAM1 in Epithelial Cell Polarization

CEACAM1 在上皮细胞极化中的作用

基本信息

批准号：
7623472
负责人：
John Ernest Shively
金额：
$ 41.64万
依托单位：
BECKMAN RESEARCH INSTITUTE/CITY OF HOPE
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-07-01 至 2011-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7623472
关键词：
ANXA2 gene Acinus organ component Actins Active Sites Adaptor Signaling Protein Adenocarcinoma Affect Amino Acid Sequence Amino Acids Annexins Apical Apoptosis Apoptotic Avidin Binding Biochemical Biological Process Breast Breast Cancer Cell Calmodulin Calpain Cancer cell line Carcinoembryonic Antigen Cell Adhesion Molecules Cell Line Cell membrane Cell-Cell Adhesion Cells Chimeric Proteins Colon Colon Carcinoma Complex Confocal Microscopy Crosslinker Cytoplasmic Tail Cytoskeleton Data Dominant-Negative Mutation Down-Regulation E-Cadherin Environment Epidermal Growth Factor Receptor Epithelial Cells Epithelium Event Fluorescence Resonance Energy Transfer Gene Silencing Genes Genitourinary system Genus Cola Gland Histone Acetylation Histone Deacetylase Human Milk Hyperplastic Polyp IRF1 gene ITIM In Vitro Incubated Individual Insulin Receptor Integrin beta Chains Interferon Type II Investigation Kidney Kinetics Lead Ligands Link Lipid Bilayers Location Lung Lysine MCF7 cell Malignant Neoplasms Malignant neoplasm of prostate Maps Mass Spectrum Analysis Measurement Measures Mediating Messenger RNA Methylation Mitochondria Modeling Molecular Mutate Mutation Pathway interactions Peptide Conformation Peptide Sequence Determination Peptides Phenotype Phosphorylation Phosphotransferases Play Premalignant Printing Process Proliferating Property Prostate Prostatic Neoplasms Protein Isoforms Proteins Proteomics RNA Splicing Receptor Signaling Recombinant Proteins Role Sequence Analysis Signal Pathway Signal Transduction Site Small Interfering RNA Staging Stretching Sulfhydryl Compounds Surface Plasmon Resonance T-Lymphocyte Testing Thermodynamics Tissues Transfection Transmembrane Domain Tropomyosin Two-Hybrid System Techniques Vesicle Yeasts adenoma base bisulfite cancer cell crosslink foot in vivo insight knockout gene link protein neoplastic cell o-Phthalaldehyde polymerization promoter protein complex response src-Family Kinases transcription factor transcription factor Sp2 tumor tumor progression

项目摘要

DESCRIPTION (provided by applicant): CEACAM1 is a homotypic cell adhesion molecule that plays a critical role in epithelial cell polarization, including lumen formation for breast and prostate epithelial cells when grown in 3D culture. Both long (72AA) and short (12 AA) cytoplasmic domain isoforms are produced by alternative mRNA splicing, with the short form predominant in most epithelial cells, and the long form predominant in T-cells. In addition, the CEACAM1 gene is silenced in most cancers, and in the case of colon cancer, it is silenced as early as pre-malignant hyperplastic polyps and adenomas. We have shown that peptides derived from the short form interact directly with actin, tropomyosin, calmodulin, and annexin 2, playing a role in interactions with the cytoskeleton, and that when phosphorylated on Thr and Ser residues, initiate a mitochondrial pathway of apoptosis, playing a role in lumen formation. In order to dissect these roles more fully, we propose to determine the hierarchal sequence of events that lead to productive interactions of the short form with the cytoskeleton, to identify the downstream kinases and adaptor proteins that lead to apoptosis by the short form, to determine the mechanism of receptor signaling inhibition by the long form, and to dissect the mechanism of gene silencing in prostate and colon cancers. To achieve the first three aims we propose biochemical and proteomic approaches that allow the direct identification of interacting proteins and the testing of these interactions in vivo in cells undergoing lumen formation using siRNA, confocal, and FRET approaches. In vivo foot printing and proteomic approaches will be used to identify the promoter complexes responsible for gene silencing in prostate and colon cell lines that do or do not express CEACAM1. Functional analyses of identified factors will be performed by factor depletion using siRNA approaches. These studies should provide a mechanistic insight into the function of CEACAM1 in a relevant biological process, namely lumen formation in normal differentiation, and the mechanism of CEACAM1 gene silencing in cancers of epithelial cell origin and the consequences thereof.

描述（由申请人提供）：CEACAM1 是一种同型细胞粘附分子，在上皮细胞极化中发挥关键作用，包括乳腺和前列腺上皮细胞在 3D 培养物中生长时的管腔形成。长 (72AA) 和短 (12 AA) 胞质结构域亚型都是通过选择性 mRNA 剪接产生的，其中短形式在大多数上皮细胞中占主导地位，而长形式在 T 细胞中占主导地位。此外，CEACAM1基因在大多数癌症中都是沉默的，就结肠癌而言，它早在癌前增生性息肉和腺瘤中就被沉默了。我们已经证明，源自短形式的肽直接与肌动蛋白、原肌球蛋白、钙调蛋白和膜联蛋白 2 相互作用，在与细胞骨架的相互作用中发挥作用，并且当 Thr 和 Ser 残基磷酸化时，启动线粒体凋亡途径，发挥作用管腔形成中的作用。为了更全面地剖析这些作用，我们建议确定导致短形式与细胞骨架产生有效相互作用的事件的层次序列，以确定短形式导致细胞凋亡的下游激酶和接头蛋白，以确定长形式抑制受体信号传导的机制，并剖析前列腺癌和结肠癌中基因沉默的机制。为了实现前三个目标，我们提出了生化和蛋白质组学方法，这些方法可以直接鉴定相互作用的蛋白质，并使用 siRNA、共聚焦和 FRET 方法在正在进行管腔形成的细胞中体内测试这些相互作用。体内足印和蛋白质组学方法将用于鉴定在表达或不表达 CEACAM1 的前列腺和结肠细胞系中负责基因沉默的启动子复合物。将使用 siRNA 方法通过因子去除来对已识别因子进行功能分析。这些研究应该为 CEACAM1 在相关生物过程中的功能提供机制上的见解，即正常分化中的管腔形成，以及上皮细胞来源的癌症中 CEACAM1 基因沉默的机制及其后果。