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：
7822777
负责人：
John Ernest Shively
金额：
$ 42.47万
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 2012-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7822777
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是一种同质细胞粘附分子，在上皮细胞极化中起关键作用，包括在3D培养中生长时乳腺和前列腺上皮细胞的管腔形成。长（72AA）和短（12个AA）细胞质结构域同工型都通过替代mRNA剪接产生，大多数上皮细胞中占主导地位，而在T细胞中占主导地位。此外，在大多数癌症中，CEACAM1基因都会沉默，就结肠癌而言，早在预防前增生性息肉和腺瘤时就会被沉默。我们已经表明，源自短形式的肽直接与肌动蛋白，肌动蛋白，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了作用，并且当在THR和SER残基上磷酸化时，启动了凋亡的线粒体途径时，在肾小管中发挥了作用。 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.为了实现前三个目标，我们提出了生化和蛋白质组学方法，这些方法允许直接鉴定相互作用的蛋白质以及使用siRNA，共聚焦和FRET方法在细胞中进行体内这些相互作用的测试。体内脚印和蛋白质组学方法将用于识别负责或不表达CEACAM1的前列腺和结肠细胞系中的基因沉默的启动子复合物。鉴定因子的功能分析将通过使用siRNA方法通过因子耗竭来进行。这些研究应提供对CEACAM1在相关生物学过程中的功能，正常分化中的管腔形成以及CEACAM1基因沉默在上皮细胞起源及其后果及其后果中的机理中的机械洞察力。