unbiased analysis of tcrαβ chains at the single-cell level in human cd8+ t-cell subsets无偏tcrαβ链分析人类cd8 + t细胞在单细胞水平的子集.pdfVIP

Unbiased Analysis of TCRa/b Chains at the Single-Cell Level in Human CD8+ T-Cell Subsets 1 1 1 1 1 Xiaoming Sun , Masumichi Saito , Yoshinori Sato , Takayuki Chikata , Takuya Naruto , 2 2 2 2 1 Tatsuhiko Ozawa , Eiji Kobayashi , Hiroyuki Kishi , Atsushi Muraguchi , Masafumi Takiguchi * 1 Center for AIDS Research, Kumamoto University, Honjo, Kumamoto, Japan, 2 Department of Immunology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan Abstract T-cell receptor (TCR) a/ b chains are expressed on the surface of CD8+ T-cells and have been implicated in antigen recognition, activation, and proliferation. However, the methods for characterization of human TCRa/ b chains have not been well established largely because of the complexity of their structures owing to the extensive genetic rearrangements that they undergo. Here we report the development of an integrated 5 9-RACE and multiplex PCR method to amplify the full- length transcripts of TCRa/ b at the single-cell level in human CD8+ subsets, including naive, central memory, early effector memory, late effector memory, and effector phenotypic cells. Using this method, with an approximately 47% and 62% of PCR success rate for TCRa and for TCRb chains, respectively, we were able to analyze more than 1,000 reads of transcripts of each TCR chain. Our comprehensive analysis revealed the following: (1) chimeric rearrangements of TCRd-a, (2) control of TCRa/ b transcription with multiple transcriptional initiation sites, (3) altered utilization of TCRa/ b chains in CD8+ subset