Nutt S L, Morrison A M, Dörfler P, Rolink A, Busslinger M
Research Institute of Molecular Pathology, Dr. Bohr-Gasse 7, A-1030 Vienna, Austria.
EMBO J. 1998 Apr 15;17(8):2319-33. doi: 10.1093/emboj/17.8.2319.
The Pax-5 gene codes for the transcription factor BSAP which is essential for the progression of adult B lymphopoiesis beyond an early progenitor (pre-BI) cell stage. Although several genes have been proposed to be regulated by BSAP, CD19 is to date the only target gene which has been genetically confirmed to depend on this transcription factor for its expression. We have now taken advantage of cultured pre-BI cells of wild-type and Pax-5 mutant bone marrow to screen a large panel of B lymphoid genes for additional BSAP target genes. Four differentially expressed genes were shown to be under the direct control of BSAP, as their expression was rapidly regulated in Pax-5-deficient pre-BI cells by a hormone-inducible BSAP-estrogen receptor fusion protein. The genes coding for the B-cell receptor component Ig-alpha (mb-1) and the transcription factors N-myc and LEF-1 are positively regulated by BSAP, while the gene coding for the cell surface protein PD-1 is efficiently repressed. Distinct regulatory mechanisms of BSAP were revealed by reconstituting Pax-5-deficient pre-BI cells with full-length BSAP or a truncated form containing only the paired domain. IL-7 signalling was able to efficiently induce the N-myc gene only in the presence of full-length BSAP, while complete restoration of CD19 synthesis was critically dependent on the BSAP protein concentration. In contrast, the expression of the mb-1 and LEF-1 genes was already reconstituted by the paired domain polypeptide lacking any transactivation function, suggesting that the DNA-binding domain of BSAP is sufficient to recruit other transcription factors to the regulatory regions of these two genes. In conclusion, these loss- and gain-of-function experiments demonstrate that BSAP regulates four newly identified target genes as a transcriptional activator, repressor or docking protein depending on the specific regulatory sequence context.
Pax-5基因编码转录因子BSAP，而BSAP对于成年B淋巴细胞生成超越早期祖细胞（前BⅠ）阶段的进程至关重要。尽管已有多种基因被认为受BSAP调控，但CD19是迄今为止唯一经遗传学证实其表达依赖于该转录因子的靶基因。我们利用野生型和Pax-5突变型骨髓培养的前BⅠ细胞，筛选了一大组B淋巴细胞基因以寻找其他BSAP靶基因。结果显示有四个差异表达基因受BSAP的直接调控，因为它们的表达在Pax-5缺陷的前BⅠ细胞中可被激素诱导的BSAP-雌激素受体融合蛋白快速调节。编码B细胞受体成分Ig-α（mb-1）的基因以及转录因子N-myc和LEF-1受BSAP正向调控，而编码细胞表面蛋白PD-1的基因则被有效抑制。通过用全长BSAP或仅含配对结构域的截短形式重建Pax-5缺陷的前BⅠ细胞，揭示了BSAP不同的调控机制。只有在存在全长BSAP的情况下，IL-7信号才能有效诱导N-myc基因，而CD19合成的完全恢复则严重依赖于BSAP蛋白浓度。相比之下，缺乏任何反式激活功能的配对结构域多肽就能重建mb-1和LEF-1基因的表达，这表明BSAP的DNA结合结构域足以将其他转录因子招募到这两个基因的调控区域。总之，这些功能缺失和功能获得实验表明，BSAP根据特定的调控序列背景，作为转录激活因子、阻遏因子或对接蛋白来调控四个新鉴定的靶基因。
