Division of Infection and Immunity, Department of Immunology and Molecular Pathology, Rayne Building, University College London, London, UK.
Genes Immun. 2013 Dec;14(8):479-86. doi: 10.1038/gene.2013.41. Epub 2013 Aug 29.
Interleukin-6 (IL-6) is an important pro-inflammatory cytokine involved in many autoimmune and inflammatory diseases. We have shown previously that a region from -5307 to -5202 bp upstream of the IL-6 transcriptional start site is responsible for basal IL-6 gene expression, and that there were DNA-binding proteins involved from electrophoretic mobility shift assay (EMSA) and transient expression experiments. Here we have combined surface plasmon resonance technology with mass spectrometry analysis and have identified nuclear proteins bound to this region. HNRNPA1 and HNRNPA2B1 were found consistently. EMSA supershift and chromatin immunoprecipitation assays confirmed the involvement of HNRNPA1, but not of HNRNPA2B1. Knocking down the HNRNPA1 expression by small interfering RNA resulted in reduced IL-6 transcriptional activity as assessed from transfection experiments using reporter constructs, mRNA and protein measurements. Overexpression of HNRNPA1 cDNA increased IL-6 mRNA expression. This regulation was dependent on the presence of the sequence from -5307 to -5202 bp of the IL-6 gene. Thus, HNRNPA1 is a novel transcriptional regulator of IL-6 expression, acting via the 5'-flanking sequence of the gene.
白细胞介素-6 (IL-6) 是一种重要的促炎细胞因子，参与许多自身免疫和炎症性疾病。我们之前已经表明，IL-6 转录起始位点上游的 -5307 到-5202bp 区域负责基础的 IL-6 基因表达，并且从电泳迁移率变动分析（EMSA）和瞬时表达实验中发现了涉及的 DNA 结合蛋白。在这里，我们将表面等离子体共振技术与质谱分析相结合，鉴定出与该区域结合的核蛋白。一致地发现了 HNRNPA1 和 HNRNPA2B1。EMSA 超迁移和染色质免疫沉淀测定证实了 HNRNPA1 的参与，但 HNRNPA2B1 则没有。通过小干扰 RNA 敲低 HNRNPA1 的表达，从转染实验使用报告基因构建体、mRNA 和蛋白质测量评估，导致 IL-6 转录活性降低。HNRNPA1 cDNA 的过表达增加了 IL-6 mRNA 的表达。这种调节依赖于 IL-6 基因从 -5307 到-5202bp 的序列的存在。因此，HNRNPA1 是 IL-6 表达的新型转录调节剂，通过基因的 5'-侧翼序列发挥作用。
