Neuberger M S
MRC Laboratory of Molecular Biology, Cambridge, UK.
EMBO J. 1983;2(8):1373-8. doi: 10.1002/j.1460-2075.1983.tb01594.x.
A plasmid including a mouse immunoglobulin mu gene was transfected into the IgG-secreting human lymphoid line HMy2 and mouse B- and pre-B-cell lines WEHI 231 and 18-81; stably transfected cells were selected. Transfected HMy2 cells synthesized mouse immunoglobulin mu chains as a major secreted protein but the WEHI 231 and 18-81 transfectants transcribed the introduced mu gene at lower levels. In HMy2 transfectants, most of the transcription of the introduced heavy chain gene initiated 40 and 62 bp upstream of the beginning of the VH exon translation start, although a small proportion of transcripts initiating further upstream was detected. WEHI 231 and 18-81 transfectants gave a much higher proportion of upstream initiation. Transient expression of the VH exon was monitored following transfection of mouse myeloma with the VH gene DNA in various plasmid constructs. VH transcription was only observed if the plasmids contained a segment derived from the large VH-CH intron of the immunoglobulin heavy chain locus. This segment, located between JH and switch regions, functioned both downstream of the VH exon and upstream in either orientation. The existence of a transcription enhancer element in this region is therefore proposed.
将包含小鼠免疫球蛋白μ基因的质粒转染到分泌IgG的人淋巴细胞系HMy2以及小鼠B细胞和前B细胞系WEHI 231和18 - 81中;筛选出稳定转染的细胞。转染后的HMy2细胞合成小鼠免疫球蛋白μ链作为主要分泌蛋白,但WEHI 231和18 - 81转染细胞中转染的μ基因转录水平较低。在HMy2转染细胞中,引入的重链基因的大部分转录起始于VH外显子翻译起始点上游40和62 bp处,不过也检测到一小部分转录本起始于更上游的位置。WEHI 231和18 - 81转染细胞中上游起始的比例要高得多。用各种质粒构建体中的VH基因DNA转染小鼠骨髓瘤后,监测VH外显子的瞬时表达。只有当质粒包含来自免疫球蛋白重链基因座的大VH - CH内含子的片段时,才会观察到VH转录。该片段位于JH和转换区之间,无论其方向如何,在VH外显子下游和上游均起作用。因此,推测该区域存在转录增强元件。