Pereira P, Gerber D, Regnault A, Huang S Y, Hermitte V, Coutinho A, Tonegawa S
Unite d'Immunobiologie CNRS URA 1961 andUnite de Biologie Moleculaire du Gene, U277 INSERM, Institut Pasteur, Paris, France.
Int Immunol. 1996 Jan;8(1):83-90. doi: 10.1093/intimm/8.1.83.
We have recently described a mAb (2.11) that recognizes the Vgamma1-Jgamma4-Cgamma4 chain. With this mAb and an anti-delta mAb we separated gammadelta+ 2.11(+) and gammadelta+ 2.11(-) intraepithelial lymphocytes (i-IEL) by FACS. Transcripts of rearranged TCR Vgamma1 and Vdelta2 genes in both i-IEL populations were analyzed by PCR followed by sequence analysis of cDNA spanning the junction of variable (V) and joining (J) genes. Roughly the same number of Vgamma1 and Vgamma2 transcripts were found in the 2.11(+) population while >90% of the transcripts in the 2.11(-)population contained a Vgamma gene sequence. Furthermore, gamma transcripts in the 2.11+ population were functional, while only 30-40% of the Vgamma transcripts in either population contained an in-frame sequence. The observed frequency of transcripts is what would be expected from cell populations that have not gone through cellular selection mediated by the RCR. Expression of Vgamma mRNA in RCRalphabeta and TCRgammadelta thymocytes was studied by a technique that analyzed populations of transcripts of rearranged genes. In both T cell population similar levels of Vgamma transcripts were found and about two out of three transcripts were out-of-frame. During the cloning and sequence analysis, we indentified a clone that expresses the Vgamma segment rearranged to the Jgamma3-Cgamma region in C57BL/6 mice. Together with the PCR cloning and sequencing of the complete Cgamma region in C57BL/6 mice these data demonstrate that the Jgamma-Cgamma gene is functional in this strain. Taken together, these studies revealed that: (i) cells expressing the Vgamma1 chain are an important subset of the gammadelta i-IEL population and that they show extensive junctional diversity; (ii) there is no correlation between expression of in-frame Vgamma transcripts and expression of Vgamma2 chains at the cell surface; and (iii) cells expressing the Vgamma3 chain might be a minor subset of the gammadelta T cell population in C57BL/6 mice.
我们最近描述了一种识别Vgamma1-Jgamma4-Cgamma4链的单克隆抗体（2.11）。利用该单克隆抗体和一种抗δ单克隆抗体，我们通过荧光激活细胞分选术（FACS）分离出γδ+ 2.11（+）和γδ+ 2.11（-）上皮内淋巴细胞（i-IEL）。通过聚合酶链反应（PCR）分析这两个i-IEL群体中重排的TCR Vgamma1和Vdelta2基因的转录本，随后对跨越可变（V）基因和连接（J）基因连接处的cDNA进行序列分析。在2.11（+）群体中发现的Vgamma1和Vgamma2转录本数量大致相同，而在2.11（-）群体中，超过90%的转录本包含Vgamma基因序列。此外，2.11+群体中的γ转录本具有功能，而两个群体中只有30 - 40%的Vgamma转录本包含读框内序列。观察到的转录本频率与未经过由RCR介导的细胞选择的细胞群体所预期的频率一致。通过一种分析重排基因转录本群体的技术，研究了RCRαβ和TCRγδ胸腺细胞中Vgamma mRNA的表达。在两个T细胞群体中发现了相似水平的Vgamma转录本，并且大约三分之二的转录本是移码的。在克隆和序列分析过程中，我们鉴定出一个在C57BL/6小鼠中表达重排至Jgamma3-Cgamma区域的Vgamma片段的克隆。连同对C57BL/6小鼠完整Cgamma区域的PCR克隆和测序，这些数据表明Jgamma-Cgamma基因在该品系中具有功能。综上所述，这些研究揭示：（i）表达Vgamma1链的细胞是γδ i-IEL群体的一个重要亚群，并且它们表现出广泛的连接多样性；（ii）读框内Vgamma转录本的表达与细胞表面Vgamma2链的表达之间没有相关性；（iii）表达Vgamma3链的细胞可能是C57BL/6小鼠中γδ T细胞群体的一个次要亚群。
