An 'environment to nucleus' signaling system operates in B lymphocytes: redox status modulates BSAP/Pax-5 activation through Ref-1 nuclear translocation.
An 'environment to nucleus' signaling system operates in B lymphocytes: redox status modulates BSAP/Pax-5 activation through Ref-1 nuclear translocation.
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DOI：
10.1093/nar/28.5.1099
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发表时间：
2000-03
影响因子：
14.9
通讯作者：
G. Tell;Alessandro Zecca;L. Pellizzari;P. Spessotto;A. Colombatti;M. Kelley;G. Damante;C. Pucillo
中科院分区：
文献类型：
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作者：
G. Tell;Alessandro Zecca;L. Pellizzari;P. Spessotto;A. Colombatti;M. Kelley;G. Damante;C. Pucillo
Ref-1（也称为APE或HAP 1）蛋白是一种影响多种重要细胞功能的双功能酶。它是DNA碱基切除修复途径的主要成员。此外，Ref-1通过还原高度反应性的半胱氨酸残基来刺激几种转录因子（TF）的DNA结合活性。因此，它代表了一种以快速方式调节真核基因表达的机制。然而，它已被证明，外部刺激直接作用于Ref-1，通过增加其表达水平，一个耗时的机制，代表了一个矛盾的TF调节的速度。在本文中，我们证明，这只是一个明显的悖论。通过蛋白质印迹分析和脉冲追踪实验评估，B淋巴细胞暴露于H（2）O（2）诱导细胞核中Ref-1蛋白水平快速和持续增加。一个时间过程，双色原位免疫细胞化学表明Ref-1在细胞核中的上调在<30 min时主要是其胞质形式移位的结果。这种早期核积累在调节B细胞特异性激活蛋白BSAP/Pax-5的DNA结合活性中是有效的。事实上，EMSA实验表明，与Ref-1的瞬时相互作用上调BSAP/Pax-5的DNA结合活性。此外，在共转染实验中，Ref-1将对CD 19启动子的寡聚化BSAP/Pax-5结合位点的BSAP/Pax-5活化作用提高了5至8倍。因此，Ref-1通过上调BSAP/Pax-5的DNA结合活性介导其作用，解释了B细胞中新的快速外部/内部信号传导途径。
The Ref-1 (also called APE or HAP1) protein is a bifunctional enzyme impacting on a wide variety of important cellular functions. It acts as a major member of the DNA base excision repair pathway. Moreover, Ref-1 stimulates the DNA-binding activity of several transcription factors (TFs) through the reduction of highly reactive cysteine residues. Therefore, it represents a mechanism that regulates eukaryotic gene expression in a fast way. However, it has been demonstrated that external stimuli directly act on Ref-1 by increasing its expression levels, a time-consuming mechanism representing a paradox in terms of rapidity of TF regulation. In this paper we demonstrate that this is only an apparent paradox. Exposure of B lymphocytes to H(2)O(2)induced a rapid and sustained increase in Ref-1 protein levels in the nucleus as evaluated by both western blot analysis and by pulse-chase experiments. A time course, two color in situ immunocytochemistry indicated that the up-regulation of Ref-1 in the nucleus at <30 min was primarily the consequence of translocation of its cytoplasmic form. This early nuclear accumulation is effective in modulating the DNA-binding activity of the B cell-specific activator protein BSAP/Pax-5. In fact, EMSA experiments demonstrate that a transient interaction with Ref-1 up-regulates the DNA-binding activity of BSAP/Pax-5. Moreover, in a co-transfection experiment, Ref-1 increased the BSAP/Pax-5 activating effect on an oligomerized BSAP/Pax-5 binding site of the CD19 promoter by 5- to 8-fold. Thus, Ref-1 mediates its effect by up-regulating the DNA-binding activity of BSAP/Pax-5, accounting for a new and fast outside/inside pathway of signaling in B cells.