Structure, function and disease susceptibility of bovine MHC class II molecules

牛白细胞抗原(BOLA)II类基因编码高度多态的跨膜糖蛋白，将抗原肽呈递给辅助T细胞。BOLA-DRB3被认为是最具功能意义的基因，因为它被活跃地转录，该基因座已被鉴定出超过102个等位基因。首先，我们开发了基于聚合酶链式反应-序列的分型技术(PCR-SBT)，它可以在DNA序列水平上分配DRB3等位基因，并可以对大量动物进行快速筛查。接下来，我们设计测定了来自日本黑牛等四个不同品种的419头动物的BOLA-DRB3等位基因外显子2的核苷酸序列。对日本短角荷斯坦牛和泽西牛的等位基因频率、BOLA-DRB3等位基因的系统发育关系以及这些群体的遗传背景进行了分析。对81个…牛的BOLA-DRB3基因外显子2进行了核苷酸序列分析更多的白血病病毒(BLV)感染的动物，包括白血病健康、持续性淋巴细胞增多症(PL)和淋巴瘤等3个独立阶段的动物，通过基于PCR测序的分型来确定。共鉴定出23个不同的BOLA-DR83等位基因，其中包括3个新等位基因。健康牛BOLA-DRB3^*14011阳性群体明显高于同一等位基因阳性个体。相比之下，BoLA-DRB3^*1601等位基因在患PL和淋巴瘤的牛中最常见。序列分析表明，在43头感染BLV但健康的牛中，大约56%携带至少一个编码DR分子β1结构域的Arg^lt；71&gt；、Glu^&lt；74&gt；、Arg^&lt；77&gt；和Val^&lt；78&gt；的BoLA-DRB3等位基因，这表明编码KERV和RERV基序的等位基因可能对肿瘤的发展具有保护作用。相比之下，在23头感染BLV的淋巴瘤牛中，约有70%携带编码Ala^&lt；74&gt；、Thr^&lt；77&gt；和Tyr^&lt；78&gt；的两个等位基因，这表明ATY/ATY基因可能与淋巴瘤的易感性有关。接下来，为了证实MHC-II类基因的多态性与淋巴瘤之间的关系，我们感染了28只绵羊。在16只患淋巴瘤的绵羊中，38%的绵羊携带编码Arg-Lys(RK)的绵羊MHC(Ola)-DRB1基因第70和71位的特异性等位基因，而在12只处于无症状阶段的绵羊中，这一比例为83%。此外，我们用BLV囊膜糖蛋白GP 51的T辅助、T细胞毒性和B细胞表位的合成抗原肽免疫RK/XX或XX/XX基因型的绵羊，最后一次免疫后两周，所有绵羊都被BLV感染的绵羊的血液攻击。携带抗性等位基因的绵羊对混合多肽表现出较高的免疫应答，未发展为白血病。较少

The bovine leukocyte antigen (BoLA)class II genes code for highly polymorphic transmembrane glycoproteins that present antigenic peptides to helper T cells. The BoLA-DRB3 is thought to be the most functionally significant as it is actively transcribed and in excess of 102 alleles have been characterized for this locus. First, we developed the technique of polymerase chain reaction-sequence based typing(PCR-SBT)which can assign DRB3 alleles in DNA sequence level and would allow fast screening of large numbers of animals. Next, we designed to determine the nucleotide sequences of exon 2 of BoLA-DRB3 alleles in a total of 419 animalks from four distinct cattle breeds such as Japanese Black. Japanese Shorthorn Holstein and Jersey by our PCR-SBT method, and the allele frequencies and the phylogenetic relationship of BoLA-DRB3 alleles and the genetic background of these populations are discussed.The nucleotide sequencing of exon 2 of the BoLA-DRB3 gene, which is highly polymorphic, of 81 bov … More ine leukemia virus(BLV)-infected animals with 3 independent stages such as an aleukemic healthy, persistent lymphocytosis(PL)and lymphoma was determined by PCR-sequenced-based typing. We identified 23 distinct BoLA-DR83 alleles, including 3 new alleles. The population of healthy cattle positive for BoLA-DRB3^*14011 was higher than the proportion-bearing individuals in 200 control cases that were positive for the same allele. By contrast, the BoLA-DRB3^*1601 alleles was found most frequently in cattle with PL and lymphoma. Sequence analysis revealed that, approximately 56 % of 43 BLV-infected but healthy cattle carry at least one BoLA-DRB3 allele encoding Arg^<71> of Lys^<71>, Glu^<74>, Arg^<77> and Val^<78> of β1 domain of DR molecule, which suggested that alleles encoding the KERV and RERV motifs might protect against tumor development. By contrast, approximately 70% of 23 BLV-infected cattle with lymphoma carry two alleles encoding Ala^<74>, Thr^<77> and Tyr^<78>, indicating that the ATY/ATY genotype might be associated with susceptibility to lymphoma. Such, these results suggest that the existence of alleles associated with resistance and susceptibility to BLV-induced leukemogenesis.Next, to confirm the association between polymorphism of MHC class II gene and lymphoma, we infected 28 sheep with BLV.Among the 16 sheep with lymphoma, 38% carried specific alleles of the ovine MHC(OLA)-DRB1 gene encoding Arg-Lys(RK)at residues 70 and 71 as compared to 83% among the 12 sheep at the asymptomatic stage. Furthermore, we immunized sheep carrying either a RK/XX genotype or a XX/XX genotype with a synthetic antigenic peptides of T-helper, T-cytotoxic, and B-cell epitopes of BLV envelope glycoprotein gp 51, and two weeks after the last immunization, all sheep were challenged with blood from a BLV-infected sheep. Sheep carrying resistant allele showed high immunoresponse against the mixed peptides and did not proceeded to leukemia. Less