白细胞介素-1β对内皮细胞中人诱导型一氧化氮合酶基因的转录调控

Transcriptional regulation of the human iNOS gene by IL-1beta in endothelial cells.

作者信息

Kolyada A Y, Madias N E

机构信息

Department of Medicine, Tufts University School of Medicine, Boston, Massachusetts, USA.

出版信息

Mol Med. 2001 May;7(5):329-43.

PMID:11474579

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC1950040/

Abstract

BACKGROUND

Vascular endothelium participates in the control of vascular tone and function via the release of nitric oxide (NO) by the endothelial-type NO synthase (eNOS). Inducible NO synthase (iNOS) expression in endothelial cells occurs in many clinical conditions following induction by lipopolysaccharide or cytokines and generates large quantities of NO that result in endothelial cell activation and dysfunction. No information exists on the transcriptional regulation of the human iNOS gene (or that of other species) in endothelial cells.

MATERIALS AND METHODS

We examined the transcriptional regulation of the human iNOS gene by interleukin-1beta (IL-1beta) in rat pulmonary microvascular endothelial cells (PVEC) by transient cotransfections of different iNOS-promoter constructs and cDNA of different transcription factors and regulatory proteins.

RESULTS

The -1034/+88 bp iNOS promoter was strongly induced by IL-1beta, the regulatory elements for such induction being localized downstream of -205 bp. Cotransfection experiments with NF-kappaB isoforms, IkappaB isoforms, and IKK mutants suggested that the NF-kappaB site at -115/-106 bp is important, but not sufficient, for induction of iNOS promoter and that the role of NF-kappaB is partially independent of its binding site. C/EBP sites within the -205/+88 bp region were shown to be responsible, along with NF-kappaB site, for induction of iNOS promoter by IL-1beta. Overexpression of C/EBPalpha, C/EBPdelta, and liver-enriched activator protein (LAP) activated the promoter, whereas overexpression of liver-enriched inhibitory protein (LIP) strongly suppressed it. C/EBPbeta (LAP and LIP isoforms) was constitutively present in PVEC and was induced (approximately 2-fold) by IL-1beta, whereas C/EBPdelta was not constitutively expressed but was strongly induced by IL-1beta. Both C/EBPbeta and C/EBPdelta participated in DNA-protein complex formation.

CONCLUSION

Both NF-kappaB and C/EBP pathways are important for the transcriptional regulation of the human iNOS gene by IL-1beta in PVEC.

摘要

背景

血管内皮通过内皮型一氧化氮合酶（eNOS）释放一氧化氮（NO）参与血管张力和功能的调控。在内皮细胞中，诱导型一氧化氮合酶（iNOS）的表达在许多临床情况下会在脂多糖或细胞因子诱导后出现，并产生大量的NO，导致内皮细胞活化和功能障碍。目前尚无关于人iNOS基因（或其他物种）在内皮细胞中转录调控的信息。

材料与方法

我们通过瞬时共转染不同的iNOS启动子构建体以及不同转录因子和调节蛋白的cDNA，研究了白细胞介素-1β（IL-1β）对大鼠肺微血管内皮细胞（PVEC）中人iNOS基因的转录调控。

结果

-1034/+88 bp的iNOS启动子被IL-1β强烈诱导，这种诱导的调控元件位于-205 bp下游。用NF-κB亚型、IκB亚型和IKK突变体进行的共转染实验表明，位于-115/-106 bp的NF-κB位点对iNOS启动子的诱导很重要，但并不充分，且NF-κB的作用部分独立于其结合位点。-205/+88 bp区域内的C/EBP位点与NF-κB位点一起，被证明是IL-1β诱导iNOS启动子的原因。C/EBPα、C/EBPδ和肝脏富集激活蛋白（LAP）的过表达激活了启动子，而肝脏富集抑制蛋白（LIP）的过表达则强烈抑制了它。C/EBPβ（LAP和LIP亚型）在PVEC中组成性存在，并被IL-1β诱导（约2倍），而C/EBPδ不是组成性表达，但被IL-1β强烈诱导。C/EBPβ和C/EBPδ都参与了DNA-蛋白质复合物的形成。