Van Ness B G, Weigert M, Coleclough C, Mather E L, Kelley D E, Perry R P
Institute for Cancer Research, Fox Chase Center, Philadelphia, Pennsylvania 19111.
Cell. 1981 Dec;27(3 Pt 2):593-602. doi: 10.1016/0092-8674(81)90401-3.
In cells of the B-lymphocyte lineage, 8.4 kb transcripts are constitutively produced from unrearranged kappa constant region (kappa 0) loci. To help elucidate the molecular basis of this phenomenon, we have determined the nucleotide sequence surrounding the site of transcriptional initiation. The kappa 0 transcripts are initiated within a unique Eco RI fragment located about 8 kb upstream from the C kappa gene. The start site is about 36 nucleotides downstream from a Hogness consensus sequence (TGTAAAT) and nearly 200 nucleotides upstream from a sequence that is similar to those encoding the signal peptides of kappa light chains. These features, which are usually found in the 5' flanking regions of kappa variable region genes, suggest that the kappa 0 initiation sequence may be an evolutionary relic of some common ancestral 5' element. In contrast, there is no discernible V kappa-encoding element in 780 nucleotides of sequence downstream from the initiation site. From pulse-chase-labeling experiments with a pre-B-cell hybridoma line and direct measurements of transcriptional activity in isolated nuclei, we have estimated that the rate of transcription of the kappa 0 locus is significantly lower than that of a rearranged V kappa-C kappa gene. This result, together with the fact that unrearranged V kappa genes are transcriptionally silent, suggests that structural features of both the V kappa and C kappa loci contribute to the overall transcriptional efficiency of a rearranged V kappa-C kappa gene. The 8.4 kb transcripts are not processed into any stable RNA products, despite the fact that they contain some apparently normal splice junctions; rather, they are degraded within the nucleus at about half the rate with which a kappa mRNA precursor is processed. Conceivably, the transcriptional activity of the kappa 0 locus might be a prerequisite for its recombinatorial activity.
在B淋巴细胞系的细胞中，未重排的κ恒定区（κ0）基因座可组成性产生8.4 kb的转录本。为了帮助阐明这一现象的分子基础，我们确定了转录起始位点周围的核苷酸序列。κ0转录本在位于Cκ基因上游约8 kb处的一个独特的Eco RI片段内起始。起始位点位于Hogness共有序列（TGTAAAT）下游约36个核苷酸处，且在与κ轻链信号肽编码序列相似的序列上游近200个核苷酸处。这些通常在κ可变区基因的5'侧翼区域发现的特征表明，κ0起始序列可能是某个共同祖先5'元件的进化遗迹。相比之下，在起始位点下游780个核苷酸的序列中没有可识别的Vκ编码元件。通过对前B细胞杂交瘤系进行脉冲追踪标记实验以及对分离细胞核中的转录活性进行直接测量，我们估计κ0基因座的转录速率明显低于重排的Vκ-Cκ基因。这一结果，连同未重排的Vκ基因转录沉默这一事实，表明Vκ和Cκ基因座的结构特征都有助于重排的Vκ-Cκ基因的整体转录效率。尽管8.4 kb的转录本包含一些明显正常的剪接接头，但它们并未被加工成任何稳定的RNA产物；相反，它们在细胞核内以κ mRNA前体加工速率的大约一半的速度被降解。可以想象，κ0基因座 的转录活性可能是其重组活性的一个先决条件。
