Identification of Novel Alleles and Structural Haplotypes of Major Histocompatibility Complex Class I and DRB Genes in Domestic Cat (Felis catus) by a Newly Developed NGS-Based Genotyping Method
Identification of Novel Alleles and Structural Haplotypes of Major Histocompatibility Complex Class I and DRB Genes in Domestic Cat (Felis catus) by a Newly Developed NGS-Based Genotyping Method
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DOI：
10.3389/fgene.2020.00750
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发表时间：
2020-07-15
影响因子：
3.7
通讯作者：
Shiina, Takashi
中科院分区：
文献类型：
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作者：
Okano, Masaharu;Miyamae, Jiro;Shiina, Takashi
主要组织相容性复合体（MHC）是编码移植和免疫调节分子的高度多态性和重复的基因组区域。尽管众所周知特定的MHC等位基因多态性和单倍型与免疫介导的疾病在遗传上相关，但是与人类和许多其他物种相比，猫MHC（猫白细胞抗原;弗拉）遗传和单倍型结构和多样性的详细信息是有限的。为了更好地了解家猫FLA-I和FLA-DRB基因座的等位基因和等位基因单倍型多样性程度和类型，本研究利用5只家猫的外周血白色细胞，通过计算机模拟编码外显子和基于NGS的扩增子测序，鉴定了6个可表达的FLA-I基因座。然后，我们使用一种新开发的基于NGS的基因分型方法，在两个家庭的20只家猫的32个FLA-I和16个FLA-DRB序列的基因型和注释。共有14个FLA-I和7个FLA-DRB被鉴定为新的多态性序列。系统发育分析将这些序列分为6个FLA-I（FLA-E/H/K，FLA-A，FLA-J，FLA-L，FLA-O和一个暂命名为FLA-E/H/K_Rec）和4个FLA-DRB（FLA-DRB 1，FLA-DRB 3，FLA-DRB 4，和FLA-DRB 5）谱系。两个猫家族的系谱分析揭示了8种不同的弗拉结构单倍型（I类- DRB），每个单倍型具有5至8个FLA-I和2至3个FLA-DRB转录位点。很明显，这8种弗拉单倍型是通过基因重复和缺失以及通过遗传重组与新多态性的积累和/或遗传而重排而产生的。这些发现对猫品种间遗传多样性分析和疾病关联研究以及兽医学研究具有重要意义。
The major histocompatibility complex (MHC) is a highly polymorphic and duplicated genomic region that encodes transplantation and immune regulatory molecules. Although it is well-known that particular MHC allelic polymorphisms and haplotypes are genetically relate to immune-mediated diseases detailed information of the cat MHC (Feline Leukocyte Antigen; FLA) genetic and haplotypic structure and diversity is limited in comparison to humans and many other species. In this study, to better understand the degree and types of allele and allelic haplotype diversity of FLA-class I (FLA-I) and FLA-DRB loci in domestic cats, we identified six expressible FLA-I loci in peripheral white blood cells byin silicoestimation of the coding exons and NGS-based amplicon sequencing using five unrelated cats. We then used a newly developed NGS-based genotyping method to genotype and annotate 32 FLA-I and 16 FLA-DRB sequences in two families of 20 domestic cats. A total of 14 FLA-I and seven FLA-DRB were identified as novel polymorphic sequences. Phylogenetic analyses grouped the sequences into six FLA-I (FLA-E/H/K, FLA-A, FLA-J, FLA-L, FLA-O and a tentatively named FLA-E/H/K_Rec) and four FLA-DRB (FLA-DRB1, FLA-DRB3, FLA-DRB4, and FLA-DRB5) lineages. Pedigree analysis of two cat families revealed eight distinct FLA structural haplotypes (Class I - DRB) with five to eight FLA-I and two to three FLA-DRB transcribed loci per haplotype. It is evident that the eight FLA haplotypes were generated by gene duplications and deletions, and rearrangements by genetic recombination with the accumulation and/or inheritance of novel polymorphisms. These findings are useful for further genetic diversity analysis and disease association studies among cat breeds and in veterinary medicine.