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CYTOKINE INDUCTION BY A 48 KD MYCOPLASMA GENE PRODUCT

48 KD 支原体基因产物诱导细胞因子

基本信息

批准号：
6555584
负责人：
ROBERT E HALL
金额：
$ 6.85万
依托单位：
MICHIGAN STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1997
资助国家：
美国
起止时间：
1997-08-01 至 2002-11-30
项目状态：
已结题

项目摘要

Hematopoietic/lymphoid cell differentiation is an area of intense investigation, and a number of cytokines and other factors have been identified which modulate growth, differentiation, and functional activity of hematopoietic and lymphoid cells. These include a number of microbial products such as LPS, TSST-1, erythrogenic toxins A and C, and several Mycoplasma products. In the process of examining tumor cell line conditioned media for cytokines and factors with differentiation activity, several years ago we identified, purified, and began characterizing a 48 kd factor termed P48. The biochemical characteristics and biologic activities of this factor include induction of monocytic differentiation and cytolytic activity in the HL-60 (human promyelocytic) cell line, growth inhibition (but not nonspecific toxicity) toward a large panel of other human and mouse cell lines, induction of secretion of IL-1 and TNF-alpha by human peripheral blood monocytes, and induction of accumulation of mRNAs for IL1- alpha, IL1-beta, lL6, and TNF-alpha in monocytes and monocyte-like cell lines. Over the past three years, we have further characterized this factor including cDNA and genomic cloning, sequencing, expression of the gene in E. coli, and purification of the biologically active recombinant protein. We have found that P48 represents a Mycoplasma fermentans gene product, and sequence analysis revealed a number of characteristics of a lipoprotein suggesting that P48 may be functionally similar to bacterial LPS. This revised grant proposal is designed to further characterize the biochemistry, molecular biology, and biology of P48 in detail including (I) characterization of the surface or cytoplasmic receptor for P48 on monocytes and monocyte-like cell lines and possible involvement of serum binding proteins; (2) examination of the biochemistry and functional significance of post-translational modification (e.g. lipid and/or carbohydrate) of the molecule; (3) examination of the mechanism of P48 induction of inflammatory cytokines with comparison to LPS; and (4) expression in eukaryotic systems including Baculovirus/insect cells and CHO cells P48 is an important immunomodulatory molecule which may be important in the pathophysiology of Mycoplasma infections and may be useful in dissecting the mechanisms involved in mammalian hematopoietic cell differentiation and cytokine biosynthesis.

造血/淋巴样细胞分化是一个强烈的领域调查发现，与一些细胞因子等因素有关已被确定为调节生长、分化和造血细胞和淋巴细胞的功能活性。这些包括许多微生物产物，例如脂多糖、TSST-1 致红毒素A和C，以及几种支原体产品。在……里面检测肿瘤细胞系条件培养液的过程细胞因子和具有分化活性的因子，多年在此之前，我们发现、提纯并开始描述一种48kd 命名为P48的因子。生物化学特性和生物学特性该因子的活性包括诱导单核细胞 HL-60(人)细胞的分化和杀伤活性早幼粒细胞系，生长抑制(但非特异性毒性)对一大批其他人类和小鼠细胞系，人外周血诱导分泌IL-1和TNF-α的研究血单核细胞，并诱导IL-1-mRNAs聚集单核细胞和单核细胞样细胞中的α、IL-1-β、LL6和TNF-α 细胞系。在过去的三年里，我们进一步对该因子进行了鉴定，包括cDNA和基因组克隆，该基因的测序、原核表达及纯化具有生物活性的重组蛋白。我们发现P48 代表发酵支原体基因产物和序列分析揭示了脂蛋白的许多特征提示P48在功能上可能与细菌内毒素相似。这项修订后的拨款建议旨在进一步说明详细介绍了P48的生物化学、分子生物学和生物学包括(I)表面或细胞质的特征单核细胞和单核细胞样细胞系上的P48受体和可能涉及血清结合蛋白；(2)检测翻译后基因的生物化学及其功能意义 (3)分子的修饰(如脂类和/或碳水化合物)； P48诱导炎症反应机制的研究细胞因子与脂多糖的比较；(4)在真核细胞中表达包括杆状病毒/昆虫细胞和CHO细胞P48在内的系统是一种重要的免疫调节分子，可能在支原体感染的病理生理学，可用于哺乳动物造血细胞参与的机制剖析分化和细胞因子的生物合成。