STUDIES ON THE DUFFY BLOOD GROUP (FY) ANTIGEN

Duffy血型系统由两个主要抗原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Fyb定义了四种表型，Fy(a-b-)、Fy(a-b)、Fy(a-b)和Fy(a-b- )。两种抗血清都不能凝集Duffy FY(a-b-)细胞， 黑人的表型。携带FY(a-b-)红细胞的黑人不会被感染 由人类间日疟原虫和猿猴间日疟原虫感染。 我们克隆了Duffy基因，并证明它编码一种糖蛋白(GP-FY) 在337个残基中。本提案的目标是：(1)确定 GP-FY的N-末端和C-末端结构域的拓扑，这将完成 要么通过免疫化学结合整个红细胞，要么由内而外 膜小泡或双标记免疫电镜法。(2) 在N-末端区域，测定必需的氨基酸 抗体和疟疾寄生虫的识别位置。这将是 对化学合成的多肽进行酶联免疫吸附试验，如下 通过构建氨基酸缺失和替换突变体并 在K562细胞中表达。(3)非红系GP-FY基因的序列测定 肾、肺、胸腺、脾、脑。(4)作者的角色塑造 免疫细胞化学，在这些组织中的细胞(S)产生GP-FY。(5) 趋化因子主要配体结合要求的确定 承认。这将通过建造一个面板来完成 突变和缺失突变及其在K562细胞中的表达。(6) 在小鼠中搜索同源FY将在一个基因组文库中完成，该基因组文库 将用含有骨髓cdna或 含有骨髓和脑cdna序列的探针。(七)学习 Duffy抗原决定簇与Rh蛋白相互作用的研究 (Fy5)取决于Rh蛋白的存在。这项研究将会完成 在转Duffy基因的K562细胞中表达。这个项目的长期目标是 确定：(1)GP-FY蛋白功能(S)；(2) 脑细胞中的Duffy样蛋白；(3)序列构象 结合了裂殖子和药物的设计，可以阻止 侵袭性；(4)GP-FY蛋白是否为转运蛋白；最后，(5) 设计小鼠基因敲除实验以确定其意义 达菲·吉恩。这项研究项目将产生重大的生物医学后果。 在设计更灵敏和可靠的试剂检测中 达菲抗原，新生儿溶血性疾病的预测，设计 防止寄生虫入侵的药物，将GP-FY的角色定义为人类 红细胞趋化因子受体与Duffy基因功能的解开。 达菲基因在各种组织中都很活跃，这些研究将 确定它在人类生物学中的相关性。

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.