Allele-specific quantification of human leukocyte antigen transcript isoforms by nanopore sequencing

Frontiers in Immunology

Immunology

Chang Liu¹  Andrew E. O. Hughes¹  Maureen C. Montgomery²  Eric T. Weimer³ 

[1] Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO, United States;Molecular Immunology Laboratory, McLendon Clinical Laboratories, University of North Carolina Hospitals, Chapel Hill, NC, United States;Molecular Immunology Laboratory, McLendon Clinical Laboratories, University of North Carolina Hospitals, Chapel Hill, NC, United States;Department of Pathology and Laboratory Medicine, University of North Carolina at Chapel Hill School of Medicine, Chapel Hill, NC, United States;

关键词: human leukocyte antigen (HLA);    nanopore sequencing;    long-read sequencing;    transcript isoforms;    allele-specific expression;   

DOI  :  10.3389/fimmu.2023.1199618

 received in 2023-04-17, accepted in 2023-07-05,  发布年份 2023

来源: Frontiers

【 摘 要 】

IntroductionWhile tens of thousands of HLA alleles have been identified by DNA sequencing, the contribution of alternative splicing to HLA diversity is not well characterized. In this study, we sought to determine if long-read sequencing could be used to accurately quantify allele-specific HLA transcripts in primary human lymphocytes.MethodscDNA libraries were prepared from peripheral blood lymphocytes from 12 donors and sequenced by nanopore long-read sequencing. HLA reads were aligned to donor-specific reference sequences based on the known type of each donor. Allele-specific exon utilization was calculated as the proportion of reads aligning to each allele containing known exons, and transcript isoforms were quantified based on patterns of exon utilization within individual reads.ResultsSplice variants were rare among class I HLA genes (median exon retention rate 99%–100%), except for several HLA-C alleles with exon 5 spliced out of up to 15% of reads. Splice variants were also rare among class II HLA genes (median exon retention rate 98%–100%), except for HLA-DQB1. Consistent with previous work, exon 5 of HLA-DQB1 was spliced out in alleles with a mutated splice acceptor site at rs28688207. Surprisingly, a 28% loss of exon 5 was also observed in HLA-DQB1 alleles with an intact splice acceptor site at rs28688207.DiscussionWe describe a simple bioinformatic workflow to quantify allele-specific expression of HLA transcript isoforms. Further studies are warranted to characterize the repertoire of HLA transcripts expressed in different cell types and tissues across diverse populations.

【 授权许可】

Unknown   
Copyright © 2023 Hughes, Montgomery, Liu and Weimer