Dul J L, Argon Y, Winkler T, ten Boekel E, Melchers F, Mårtensson I L
Department of Pathology, University of Chicago, USA.
Eur J Immunol. 1996 Apr;26(4):906-13. doi: 10.1002/eji.1830260428.
The surrogate light chain is composed of two polypeptides, VpreB and lambda 5. In the mouse there are two VpreB genes which are 99% identical within the coding regions. Extensive restriction enzyme mapping and sequencing of these two genes showed that only the coding region and immediate 5' and 3' flanking sequences exhibited such high homology. More distal sequences have diverged considerably. The region 5' of the respective gene directed transcription of a reporter gene in a pre-B cell line, indicating that it contained promoter, and perhaps enhancer function. The VpreB2 gene is functional, as it directed the production in COS cells of a 16-kDa protein that assembled with lambda 5 and was recognized by a VpreB-specific monoclonal antibody. Using transfected COS cells expressing either VpreB1 or VpreB2, a PCR assay was developed to examine the steady state level of transcripts from each gene. When this assay was applied to a number of cell lines representing early stages of B cell differentiation, co-expression of the two genes was observed in every case. VpreB1 and VpreB2 were co-expressed in the fetal liver of CB17 mice, where peak expression of each gene occurred at days 16-17 of gestation. Similarly, adult bone marrow from several strains of mice expressed both genes. In sorted bone marrow cells expression of both VpreB genes was detected in pro-B/pre-BI and large pre-BII cells, while the RNA steady state levels were at least 100-fold lower in small pre-BII and immature/mature B cells. Finally, single-cell reverse transcriptase-polymerase chain reaction on such sorted bone marrow cells detected VpreB1 and VpreB2 expression in at least 30% of all pro-B/pre-BI cells and large Ig heavy chain, surrogate light chain (pre-B receptor) expressing pre-BII cells. These results demonstrate that the control of expression of the two VpreB genes overlaps during development. They suggest that both VpreB1 and VpreB2 polypeptides can assemble with lambda 5 and mu to form pre-B cell receptor complexes.
替代轻链由两种多肽VpreB和λ5组成。在小鼠中，有两个VpreB基因，其编码区域内的同源性为99%。对这两个基因进行广泛的限制性内切酶图谱分析和测序表明，只有编码区域以及紧邻的5'和3'侧翼序列表现出如此高的同源性。更远端的序列则有很大差异。各自基因的5'区域在一个前B细胞系中指导报告基因的转录，表明它含有启动子，可能还有增强子功能。VpreB2基因是有功能的，因为它在COS细胞中指导产生一种16 kDa的蛋白质，该蛋白质与λ5组装并被一种VpreB特异性单克隆抗体识别。利用表达VpreB1或VpreB2的转染COS细胞，开发了一种PCR检测方法来检测每个基因转录本的稳态水平。当将该检测方法应用于代表B细胞分化早期阶段的多种细胞系时，在每种情况下都观察到了这两个基因的共表达。VpreB1和VpreB2在CB17小鼠的胎肝中共表达，每个基因的表达高峰出现在妊娠第16 - 17天。同样，来自多个品系小鼠的成年骨髓也表达这两个基因。在分选的骨髓细胞中，在原B/前B I和大前B II细胞中检测到了两个VpreB基因的表达，而在小前B II和未成熟/成熟B细胞中RNA稳态水平至少低100倍。最后，对这种分选的骨髓细胞进行单细胞逆转录聚合酶链反应，在所有原B/前B I细胞和表达大Ig重链、替代轻链（前B细胞受体）的前B II细胞中，至少30%检测到了VpreB1和VpreB2的表达。这些结果表明，在发育过程中，两个VpreB基因的表达调控存在重叠。它们表明VpreB1和VpreB2多肽都可以与λ5和μ组装形成前B细胞受体复合物。
