Andresen Lars, Jensen Helle, Pedersen Marianne T, Hansen Karen A, Skov Søren
Department of Immunology, Institute of International Health, Immunology and Microbiology, University of Copenhagen, Copenhagen, Denmark.
J Immunol. 2007 Dec 15;179(12):8235-42. doi: 10.4049/jimmunol.179.12.8235.
In this study, we characterize the molecular signal pathways that lead to MHC class I chain-related protein A (MICA) expression after histone deacetylase (HDAC)-inhibitor (HDAC-i) treatment of Jurkat T cells. Chelating calcium with BAPTA-AM or EGTA potently inhibited HDAC- and CMV-mediated MICA/B expression. It was further observed that endoplasmic reticulum calcium stores were depleted after HDAC treatment. NF-kappaB activity can be induced by HDAC treatment. However, nuclear translocation of NF-kappaB p65 was not observed after HDAC treatment of Jurkat T cells and even though we could effectively inhibit p65 expression by siRNA, it did not modify MICA/B expression. To identify important elements in MICA regulation, we made a promoter construct consisting of approximately 3 kb of the proximal MICA promoter in front of GFP. Deletion analysis showed that a germinal center-box containing a putative Sp1 site from position -113 to -93 relative to the mRNA start site was important for HDAC and CMV-induced promoter activity. Sp1 was subsequently shown to be important, as targeted mutation of the Sp1 binding sequence or siRNA mediated down modulation of Sp1-inhibited MICA promoter activity and surface-expression.
在本研究中,我们对组蛋白去乙酰化酶(HDAC)抑制剂(HDAC-i)处理Jurkat T细胞后导致MHC I类链相关蛋白A(MICA)表达的分子信号通路进行了表征。用BAPTA-AM或EGTA螯合钙可有效抑制HDAC和巨细胞病毒(CMV)介导的MICA/B表达。进一步观察到,HDAC处理后内质网钙储存被耗尽。HDAC处理可诱导核因子κB(NF-κB)活性。然而,HDAC处理Jurkat T细胞后未观察到NF-κB p65的核转位,并且即使我们可以通过小干扰RNA(siRNA)有效抑制p65表达,它也不会改变MICA/B表达。为了确定MICA调控中的重要元件,我们构建了一个启动子构建体,其由相对于mRNA起始位点从-113至-93位包含推定的Sp1位点的约3 kb近端MICA启动子组成。缺失分析表明,一个生发中心框对于HDAC和CMV诱导的启动子活性很重要,该框包含相对于mRNA起始位点从-113至-93位的一个推定的Sp1位点。随后发现Sp1很重要,因为Sp1结合序列的靶向突变或siRNA介导的Sp1下调抑制了MICA启动子活性和表面表达。