Faculty of Biological Sciences, Department of Biochemistry and Molecular Biology, Universidad de Concepcion, Santiago, Chile.
J Cell Physiol. 2012 Apr;227(4):1645-52. doi: 10.1002/jcp.22882.
Recent evidence indicates that transcription factor Runx1 modulates the expression of several phenotypic markers in dorsal root ganglia (DRGs) neurons, including the pain-related P2X3 receptor. In several cell lineages C/EBP transcription factors interact with the Runx factor family members to jointly bind and activate transcription of target genes. Here, we examine whether these two transcription factors directly regulate P2X3 gene expression. Through in silico analyses of the first 2 kb of the P2X3 gene promoter we identified putative consensus-binding sites for both Runx1 and C/EBPβ transcription factors. Transient over-expression in PC12 cells of either Runx1 or C/EBPβ increases P2X3 gene promoter activity and co-expression of both factors results in an additive stimulatory effect on the promoter function. Accordingly, chromatin immunoprecipitation assays demonstrate that both Runx1 and C/EBPβ bind to the P2X3 promoter in PC12 cells expressing this gene. Site-directed mutagenesis of the proximal Runx1 and C/EBPβ consensus elements in the P2X3 promoter decrease Runx1- and C/EBPβ-mediated transcriptional activity. Moreover, C/EBPβ-mediated enhancement of the P2X3 promoter requires a functional Runx1 binding site. Altogether our results support a functional and coordinated role for Runx1 and C/EBPβ transcription factors during activation of P2X3 gene transcription.
最近的证据表明，转录因子 Runx1 调节背根神经节 (DRG) 神经元中几种表型标志物的表达，包括与疼痛相关的 P2X3 受体。在几种细胞谱系中，C/EBP 转录因子与 Runx 因子家族成员相互作用，共同结合并激活靶基因的转录。在这里，我们研究这两个转录因子是否直接调节 P2X3 基因的表达。通过对 P2X3 基因启动子的前 2kb 进行计算机分析，我们确定了 Runx1 和 C/EBPβ 转录因子的潜在共有结合位点。在 PC12 细胞中转染 Runx1 或 C/EBPβ 均可增加 P2X3 基因启动子的活性，并且这两种因子的共表达对启动子功能具有累加的刺激作用。相应地，染色质免疫沉淀分析表明，在表达该基因的 PC12 细胞中，Runx1 和 C/EBPβ 均与 P2X3 启动子结合。P2X3 启动子中近端 Runx1 和 C/EBPβ 共有结合元件的定点突变降低了 Runx1 和 C/EBPβ 介导的转录活性。此外，C/EBPβ 介导的 P2X3 启动子增强需要功能性 Runx1 结合位点。总之，我们的结果支持 Runx1 和 C/EBPβ 转录因子在 P2X3 基因转录激活过程中具有功能协调作用。
