CLONING THE GENE FOR A NOVEL TPA-INDUCED PROTEIN

我们最近鉴定并部分提纯了一种似乎是 独特的质膜蛋白，“大麻”。大麻是在……的表面发现的 正常的人类嗜酸性粒细胞、巨核细胞和血小板，但不是 在其他正常造血细胞或冰冻切片中可检测到 正常的成人组织。大麻是一种19kD的蛋白质，是一种后 通过脂类的共价键进行翻译修饰以产生成熟的 表观相对分子质量为21kd。一株大麻特异性单抗的结合 血小板上的抗大麻抗体会引起聚集和ATP释放， 提示大麻可能是信号转导中的辅助蛋白。 来自表面感受器。初步研究还表明，谨慎的 大麻的特性及其合成的调节 提高对肿瘤发生及其相互作用的理解的潜力 转化细胞和肿瘤促进剂之间的关系。 我们发现，在培养中，接触肿瘤促进剂，12-0- 十四酰佛波醇13-乙酸酯(TPA)诱导胚胎细胞表达大麻 急性单核细胞白血病患者的骨髓细胞和原始细胞 几个人类肿瘤细胞系。TPA不能通过以下方式诱导大麻的表达 循环中的白细胞正常。这表明对大麻的调节 胎儿和成人造血细胞中的基因不同，并在 肿瘤细胞。大麻的表达也可能与 某些人肿瘤细胞在裸鼠体内生长的能力，即培养的细胞 由人体细胞系在裸鼠体内形成的肿瘤 可诱导大麻合成或含有大麻(+)亚群给出 上升到身体上表达大麻的线条。 我们近期的目标是：1.提纯大麻，用于多肽分析和 单特异性和单克隆抗体的制备。多肽序列 将用于制备核苷酸探针和/或 CDNA克隆身份的确认。2.克隆鸡传染性支气管炎病毒S 大麻，DNA序列分析将允许测定大麻的氨基酸 ITS基因的序列、组织及其可能的鉴定 启动子和调控站点。它也可能给出大麻的线索 功能。3.对大麻基因调控位点进行功能定位(S)。 从大麻的特性和基因(S)获得的信息 编码它将成为未来旨在阐明 肿瘤促进剂在调控大麻基因和确定大麻基因中的作用 在正常细胞和转化细胞中发挥作用。

We have recently characterized and partially purified what appears to be a unique plasma membrane protein, "HEMP". HEMP is found on the surface of normal human eosinophils, megakaryocytes and platelets, but is not detectable on other normal hematopoietic cells or cells in frozen sections of normal adult tissues. HEMP is a 19 kD protein that is post- translationally modified by covalent linkage of lipid to yield a mature form with an apparent MW of 21 kd. Binding of a HEMP-specific monoclonal antibody to HEMP on platelets causes aggregation and ATP release, suggesting that HEMP may be an accessory protein in signal transduction from surface receptors. Initial studies also suggest that a careful characterization of HEMP and the regulation of its synthesis has the potential to enhance understanding of tumorigenesis and the interactions between transformed cells and tumor promoters. We have found that exposure, in culture, to the tumor promoter, 12-0- tetradecanoyl phorbol 13-acetate (TPA), induces expression of HEMP by fetal marrow cells, blast cells of patients with acute monocytic leukemia and several human tumor cell lines. TPA does not induce HEMP expression by normal circulating leukocytes. This indicates that regulation of the HEMP gene is different in fetal and adult hematopoietic cells and is altered in neoplastic cells. Expression of HEMP may also be associated with the ability of some human tumor cells to grow in nude mice, i.e. cells cultured from tumors formed in nude mice by human cell lines that were either inducible for HEMP synthesis or contained a HEMP (+) subpopulation gave rise to lines that express HEMP constitutively. Our immediate goals are: 1. To purify HEMP for peptide analysis and preparation of monospecific and monoclonal antibodies. Peptide sequences will be used either for preparation of nucleotide probes and/or confirmation of cDNA clone identity. 2. To clone the cDNA and gene(s) for HEMP, DNA sequence analysis will allow determination of HEMP's amino acid sequence, the organization of its gene and identification of putative promoter and regulatory sites. It may also give clues as to HEMP's function. 3. To functionally map regulatory sites in the HEMP gene(s). Information derived from characterization of HEMP and the gene(s) that encode it will be the basis for future studies aimed at elucidating the role of tumor promoters in regulating the HEMP gene and determining HEMP's function in normal and transformed cells.