STUDIES ON THE DUFFY BLOOD GROUP (FY) ANTIGEN

达菲血型（FY）抗原的研究

• 批准号: 6110460
• 负责人: A O POGO
• 金额: $ 23.65万
• 依托单位: NEW YORK BLOOD CENTER
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-01-01 至 1999-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6110460
• 关键词: blood group antigens; chemical binding; cytokine receptors; enzyme linked immunosorbent assay; erythrocyte membrane; gene expression; gene targeting; genetic mapping; genetically modified animals; human tissue; immunocytochemistry; immunoelectron microscopy; laboratory mouse; laboratory rabbit; malaria; messenger RNA; molecular shape; northern blottings; nucleic acid hybridization; nucleic acid sequence; organ; polymerase chain reaction; protein structure function; protozoal antigen; surface antigens

The Duffy blood group system consists of two major antigens Fya and Fyb produced by FY+A and FY+B co-dominant alleles. Antisera, anti-Fya and anti-Fyb define four phenotypes, Fy(a+b-), Fy(a-b+), Fy(a+b+), and Fy(a-b- ). Neither antiserum agglutinates Duffy Fy(a-b-) cells, the predominant phenotype in Blacks. Blacks with Fy(a-b-) erythrocytes cannot be infected by the human malarial parasite P. vivax and simian parasite P. knowlesi. We cloned the Duffy gene and showed that it encodes a glycoprotein (gp-Fy) of 337 residues. The objectives of this proposal are: (1) To determine the topology of the N- and C-terminal domains of gp-Fy, which will be done either by immunochemical binding of whole erythrocytes and inside-out membrane vesicles or double labeling immunoelectromicroscopy. (2) The determination, at the N-terminal domain, of the amino acids necessary for recognition sites of antibodies and malarial parasites. This will be carried out with ELISA assay of chemically synthesized peptides, followed by the construction of amino acid deletion and substitution mutants and expression in K562 cells. (3) The sequencing of non-erythroid gp-Fy mRNA in kidney, lung, thymus, spleen and brain. (4) The characterization by immunocytochemistry, of which cell(s) in these tissues produce gp-Fy. (5) Identification of the major ligand-binding requirements for chemokine recognition. This will be done by the construction of a panel of substitution and deletion mutants and expression in K562 cells. (6) The search for homologous FY in mouse will be done in a genomic library that will be screened with a probe containing either bone marrow cDNA or a probe having bone marrow and brain cDNA sequences. (7) To study interactions of Duffy and Rh protein since a Duffy antigenic determinant (Fy5) depends upon the presence of the Rh protein. The study will be done in Duffy-transfected K562 cells. The long-term goals of this project are to determine: (1) the gp-Fy protein function(s); (2) the significance of the Duffy-like protein in brain cells; (3) the sequence conformation that binds to the merozoites and the design of drugs which would block invasion; (4) whether gp-Fy protein is a transporter and finally, (5) the design of mouse knockout experiments to determine the significance of Duffy gene. This research project will have major biomedical consequences in the design of more sensitive and reliable reagents for the detection of Duffy antigens, anticipation of hemolytic diseases in the newborn, design of drugs to prevent parasite invasion, defining gp-Fy role as the human erythrocyte chemokine receptor and unraveling the function of Duffy gene. The Duffy gene is active in a variety of tissues, and these studies will establish its relevance in human biology.

达菲血型系统由两种主要抗原Fya和Fyb组成 由FY+A和FY+B共显性等位基因产生。 抗血清，抗Fya和 抗Fy B定义了四种表型，Fy（a+B-）、Fy（a-B+）、Fy（a+B+）和Fy（a-B- ). 这两种抗血清都不凝集Duffy Fy（a-b-）细胞，主要是 在黑人中的表型。 具有Fy（a-b-）红细胞的黑人不能被感染 由人类疟原虫间日疟原虫和猿类疟原虫诺氏疟原虫感染。 我们克隆了Duffy基因，并表明它编码一种糖蛋白（gp-Fy）， 337个残基。 本提案的目标是：（1）确定 gp-Fy的N-和C-末端结构域的拓扑结构，这将在 或者通过免疫化学结合整个红细胞， 膜囊泡或双标记免疫电镜。(2)的 在N-末端结构域，确定 抗体和疟疾寄生虫的识别位点。 这将是 用化学合成肽的ELISA测定进行，然后 通过构建氨基酸缺失和置换突变体， 在K562细胞中表达。(3)非红系gp-Fy mRNA的序列测定 肾脏、肺、胸腺、脾脏和大脑。(4)特征是 免疫细胞化学，这些组织中的细胞产生gp-Fy。（五） 趋化因子主要配体结合要求的鉴定 识别. 这将通过建造一个 取代和缺失突变体以及在K562细胞中的表达。(6)的 在小鼠中寻找同源FY将在基因组文库中进行， 将用含有骨髓cDNA或 具有骨髓和脑cDNA序列的探针。(7)研究 Duffy抗原决定簇与Rh蛋白的相互作用 (Fy5)取决于Rh蛋白的存在。 这项研究将在 在Duffy转染的K562细胞中。 该项目的长期目标是 为了确定：（1）gp-Fy蛋白功能;（2） 脑细胞中的Duffy样蛋白;（3） 与裂殖子结合， （4）gp-Fy蛋白是否是一种转运蛋白，以及（5） 设计小鼠基因敲除实验，以确定 达菲·吉恩 这项研究项目将产生重大的生物医学影响 在设计更灵敏和可靠的检测试剂时， Duffy抗原，新生儿溶血性疾病的预测，设计 的药物，以防止寄生虫入侵，定义gp-Fy的作用，作为人类 红细胞趋化因子受体和Duffy基因功能的研究。 达菲基因在各种组织中都很活跃，这些研究将 在人类生物学中建立其相关性。