Lipkowitz S, Stern M H, Kirsch I R
Navy Medical Oncology Branch, Naval Hospital, National Cancer Institute, Bethesda, Maryland.
J Exp Med. 1990 Aug 1;172(2):409-18. doi: 10.1084/jem.172.2.409.
In this paper, using polymerase chain reaction (PCR), we demonstrated the occurrence of hybrid genes formed by interlocus recombination between T cell receptor gamma (TCR-gamma) variable (V) regions and TCR-beta joining (J) regions in the peripheral blood lymphocytes (PBL) from normal individuals and patients with ataxia-telangiectasia (AT). Sequence analysis of the PCR-derived hybrid genes confirmed that site-specific V gamma-J beta recombination had occurred and showed that 10 of 23 genomic hybrid genes maintained a correct open reading frame. By dilution analysis, the frequency of these hybrid genes was 8 +/- 1/10(5) cells in normal PBL and 587 +/- 195/10(5) cells in AT PBL. These frequencies and the approximately 70-fold difference between the normal and AT samples are consistent with previous cytogenetic data examining the occurrence of an inversion of chromosome 7 in normal and AT PBL. We also demonstrated expression of these hybrid genes by PCR analysis of first-strand cDNA prepared from both normal and AT PBL. Sequence analysis of the PCR-amplified transcripts showed that, in contrast to the genomic hybrid genes, 19 of 22 expressed genes maintained a correct open reading frame at the V-J junction and correctly spliced the hybrid V-J exon to a TCR-beta constant region, thus allowing translation into a potentially functional hybrid TCR protein. Another type of hybrid TCR transcript was found in a which a rearranged TCR-gamma V-J exon was correctly spliced to a TCR-beta constant region. This form of hybrid gene may be formed by trans-splicing. These hybrid TCR genes may serve to increase the repertoire of the immune response. In addition, studies of their mechanism of formation and its misregulation in AT may provide insight into the nature of the chromosomal instability syndrome associated with AT. The mechanism underlying hybrid gene formation may be analogous to the mechanism underlying rearrangements between putative growth-affecting genes and the antigen receptor loci, which are associated with AT lymphocyte clones and lymphoid malignancies.
在本文中，我们使用聚合酶链反应（PCR）证明了在正常个体和共济失调毛细血管扩张症（AT）患者的外周血淋巴细胞（PBL）中，T细胞受体γ（TCR-γ）可变（V）区与TCR-β连接（J）区之间通过基因座间重组形成了杂交基因。对PCR衍生的杂交基因进行序列分析证实，发生了位点特异性Vγ-Jβ重组，并表明23个基因组杂交基因中有10个保持了正确的开放阅读框。通过稀释分析，这些杂交基因在正常PBL中的频率为8±1/10⁵细胞，在AT PBL中的频率为587±195/10⁵细胞。这些频率以及正常样本与AT样本之间约70倍的差异与先前检查正常和AT PBL中7号染色体倒位发生情况的细胞遗传学数据一致。我们还通过对从正常和AT PBL制备的第一链cDNA进行PCR分析，证明了这些杂交基因的表达。对PCR扩增转录本的序列分析表明，与基因组杂交基因相反，22个表达基因中有19个在V-J连接处保持了正确的开放阅读框，并将杂交V-J外显子正确剪接到TCR-β恒定区，从而允许翻译成潜在的功能性杂交TCR蛋白。在另一种情况中发现了另一种类型的杂交TCR转录本，其中重排的TCR-γ V-J外显子被正确剪接到TCR-β恒定区。这种杂交基因形式可能是通过反式剪接形成的。这些杂交TCR基因可能有助于增加免疫反应的多样性。此外，对它们在AT中的形成机制及其调控异常的研究可能有助于深入了解与AT相关的染色体不稳定综合征的本质。杂交基因形成的潜在机制可能类似于假定的生长影响基因与抗原受体基因座之间重排的潜在机制，这些重排与AT淋巴细胞克隆和淋巴恶性肿瘤有关。
