Transcriptional regulation of the murine 1-cys peroxiredoxin gene by the B cell-specific activator protein, Pax5
Transcriptional regulation of the murine 1-cys peroxiredoxin gene by the B cell-specific activator protein, Pax5
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DOI：
10.1002/jcb.21638
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发表时间：
2008-05-15
影响因子：
4
通讯作者：
Pak, Jhang Ho
中科院分区：
文献类型：
--
作者：
Lee, In-Seon;Choi, Won Ho;Pak, Jhang Ho
Pax 5是一种B细胞特异性激活蛋白（13 SAP），属于转录因子的配对盒基因家族，在控制B淋巴细胞生成过程中谱系和分化阶段特异性基因的表达中起重要作用。我们确定了两个假定的Pax 5结合位点在668 bp的小鼠1-cys peroxiredoxin（1-cysPrx）启动子区。这些位点位于距离翻译起始位点-278至-262和-50至-34的位置。凝胶迁移率变动分析表明，重组Pax 5蛋白特异性地结合到核苷酸区域-56至-24（MPI探针）和-284至-253（MP2探针）。此外，来自B淋巴母细胞（IM-9）的内源性Pax 5蛋白与MP 1和MP 2探针形成DNA-蛋白复合物，表明Pax 5结合特异性地发生在体内这些序列上。瞬时转染研究表明，外源Pax 5的表达导致1-cysPrx启动子活性的剂量依赖性增加，这意味着Pax 5作为1-cysPrx表达的正转录因子发挥作用。进一步的瞬时共转染研究表明，Pax 5与组蛋白乙酰转移酶（HATs）如p300、cAMP反应元件结合蛋白（CBP）和p300/CBP相关因子（PCAF）的共表达增强了Pax 5介导的1-cysPrx启动子活性。免疫沉淀研究表明Pax 5直接与HAT结合。染色质免疫沉淀分析表明，Pax 5的启动子的招聘诱导组蛋白H3和H4的染色质的超乙酰化。脂多糖（LPS）刺激小鼠脾细胞导致Pax 5和1-cysPrx蛋白的协调表达。这些发现表明Pax 5可能作为1-cysPrx基因表达的反式激活因子发挥作用。
Pax5, a member of the paired box gene family of transcription factors, is a B cell-specific activator protein (13 SAP) that plays important roles in controlling the expression of lineage- and differentiation-stage specific genes during B lymphopoiesis. We identified two putative Pax5 binding sites in a 668 bp of the murine 1-cys peroxiredoxin (1-cysPrx) promoter region. These sites were located at positions -278 to -262 and -50 to -34 from the translation start site. Gel mobility shift assays showed that recombinant Pax5 protein bound specifically to the nucleotide regions -56 to -24 (MPI probe) and -284 to -253 (MP2 probe). Furthermore, endogenous Pax5 protein from B lymphoblast cells (IM-9) formed a DNA-protein complex with MP1 and MP2 probes, indicating that Pax5 binding occurs specifically at these sequences in vivo. Transient transfection studies showed that expression of exogenous Pax5 resulted in dose-dependent increases in 1-cysPrx promoter activity, implying that Pax5 functions as a positive transcription factor for 1-cysPrx expression. Further transient cotransfection studies showed that coexpression of Pax5 and histone acetyltransferases (HATs), such as p300, cAMP-response-element-binding protein (CBP) and p300/CBP associated factor (PCAF) enhanced the Pax5-mediated 1-cysPrx promoter activity. Immunoprecipitation studies indicated that Pax5 directly binds to HATs. Chromatin immunoprecipitation assays showed that the recruitment of Pax5 to the promoter induced histone H3 and H4 hyperacetylation of chromatin. Lipopolysaccharides (LPS) stimulation of murine splenocytes resulted in coordinated expression of Pax5 and 1-cysPrx proteins. These findings suggest that Pax5 may function as a transactivator of 1-cysPrx gene expression.