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首页> 外文期刊>Proteomics >Proteomic analysis of nuclear proteins from proliferative and differentiated human colonic intestinal epithelial cells
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Proteomic analysis of nuclear proteins from proliferative and differentiated human colonic intestinal epithelial cells

机译：增殖和分化的人结肠小肠上皮细胞中核蛋白的蛋白质组学分析

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摘要

Self-renewing tissues such as the intestine contain progenitor proliferating cells which subsequently differentiate. Cell proliferation and differentiation involve gene regulation processes which take place in the nucleus. A human intestinal epithelial cell line model (Caco2/TC7) which reproduces these dynamic processes has been used to perform proteomic studies on nuclear proteins. Nuclei from Caco2/TC7 cells at proliferative and differentiated stages were purified by subcellular fractionation. After two-dimensional gel electrophoresis separation and ruthenium staining, 400 protein spots were detected by image analysis. Eighty-five spots corresponding to 60 different proteins were identified by matrix-assisted laser desorption/ionization mass spectrometry in nuclei from proliferative cells. Comparison of nuclear proteomes from proliferative or differentiated cells by differential display resulted in the identification of differentially expressed proteins such as nucleolin, hnRNP A2/B1 and hnRNP A1. By using Western blot analysis, we found that the expression and number of specific isoforms of these nuclear proteins decreased in differentiated cells. Immunocytochemistry experiments also showed that in proliferative cells nucleolin was distributed in nucleoli-like bodies. In contrast, hnRNPs A2/B1 and A1 were dispersed throughout the nucleus. This study of the nuclear proteome from intestinal epithelial cells represents the first step towards the establishment of a protein database which will be a valuable resource in future studies on the differential expression of nuclear proteins in response to physiological, pharmacological and pathological modulations.
机译：自我更新的组织（例如肠道）包含祖细胞增殖细胞，这些细胞随后会分化。细胞增殖和分化涉及发生在细胞核中的基因调节过程。复制这些动态过程的人类肠上皮细胞系模型（Caco2 / TC7）已用于对核蛋白进行蛋白质组学研究。通过亚细胞分级分离纯化来自Caco2 / TC7细胞处于增殖和分化阶段的细胞核。二维凝胶电泳分离和钌染色后，通过图像分析检测到400个蛋白质斑点。通过基质辅助激光解吸/电离质谱法在增殖细胞核中鉴定出与60种不同蛋白质相对应的85个斑点。通过差异展示比较来自增殖或分化细胞的核蛋白质组，鉴定出差异表达的蛋白质，如核仁蛋白，hnRNP A2 / B1和hnRNP A1。通过使用蛋白质印迹分析，我们发现这些核蛋白在分化细胞中的表达和数量有所减少。免疫细胞化学实验还表明，核仁素在增殖细胞中分布在核仁样体中。相反，hnRNP A2 / B1和A1分散在整个核中。肠道上皮细胞核蛋白质组的这项研究代表了建立蛋白质数据库的第一步，这将是未来研究响应于生理，药理和病理学调节的核蛋白质差异表达的宝贵资源。

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