Fulton J E, Thacker E L, Bacon L D, Hunt H D
USDA-Agricultural Research Service, Avian Disease and Oncology Laboratory, East Lansing, MI 48823, USA.
Eur J Immunol. 1995 Jul;25(7):2069-76. doi: 10.1002/eji.1830250740.
Similarities between the physical structures of avian and mammalian major histocompatibility complex (MHC) class I glycoproteins have been proposed based on comparative alignment of their amino acid sequences. To investigate the physical structure of the chicken class I glycoprotein, we cloned the cDNA representing the BFIV locus of the B21 haplotype. A unique, chimeric class I glycoprotein was constructed by incorporating an epitope tag (FLAG) at the N terminus. Monoclonal antibodies to the FLAG epitope served to monitor cell-surface expression for functional analysis of the BFIV21 class I glycoprotein. The chimeric class I glycoprotein was expressed in target cells using an avian leukosis virus (ALV)-derived retrovirus vector (RCASBP). The presence of the FLAG epitope did not interfere with either alloantibody recognition or cytotoxic T lymphocyte interaction. Functional analysis employing site-directed mutagenesis identified BF amino acid residues forming serologic epitopes as well as residues important in antigen presentation to ALV-induced cytotoxic T lymphocytes. BF residues 78 and 81, corresponding to HLA 79 and 82, form an antibody epitope with a slight effect on ALV antigen presentation, consistent with their predicted orientation based on the HLA-A2 crystal structure. Alignment of the BFIV21 sequence with previously published BFIV sequences revealed polymorphisms at position 34 (HLA 34), a monomorphic residues in HLA and H-2. Residue 34 is located in pocket B and is predicted to contact the main-chain carbon of peptides bound in HLA-A2. A site-directed substitution in BFIV residue 34 dramatically alters ALV antigen presentation by the BFIV21 class I glycoprotein. These data indicate that the physical molecular structure of the chicken MHC class I glycoprotein is similar to HLA.
基于禽类和哺乳动物主要组织相容性复合体（MHC）I类糖蛋白氨基酸序列的比较比对，人们提出了它们物理结构上的相似性。为了研究鸡I类糖蛋白的物理结构，我们克隆了代表B21单倍型BFIV基因座的cDNA。通过在N端掺入一个表位标签（FLAG）构建了一种独特的嵌合I类糖蛋白。针对FLAG表位的单克隆抗体用于监测细胞表面表达，以对BFIV21 I类糖蛋白进行功能分析。使用禽白血病病毒（ALV）衍生的逆转录病毒载体（RCASBP）在靶细胞中表达嵌合I类糖蛋白。FLAG表位的存在既不干扰同种抗体识别，也不干扰细胞毒性T淋巴细胞相互作用。采用定点诱变的功能分析确定了形成血清学表位的BF氨基酸残基以及在向ALV诱导的细胞毒性T淋巴细胞呈递抗原中起重要作用的残基。与HLA 79和82相对应的BF残基78和81形成一个抗体表位，对ALV抗原呈递有轻微影响，这与基于HLA - A2晶体结构预测的方向一致。BFIV21序列与先前发表的BFIV序列比对显示，在位置34（HLA 34）存在多态性，而该位置在HLA和H - 2中是单态残基。残基34位于口袋B中，预计会与结合在HLA - A2中的肽的主链碳接触。BFIV残基34的定点取代显著改变了BFIV21 I类糖蛋白的ALV抗原呈递。这些数据表明鸡MHC I类糖蛋白的物理分子结构与HLA相似。
