Johnson D R, Pober J S
Boyer Center for Molecular Medicine, Yale Medical School, New Haven, Connecticut 06536-0812.
Mol Cell Biol. 1994 Feb;14(2):1322-32. doi: 10.1128/mcb.14.2.1322-1332.1994.
The cytokines tumor necrosis factor (TNF), beta interferon (IFN-beta), and IFN-gamma increase major histocompatibility complex class I molecule expression. A greater than additive (i.e., synergistic) induction of class I heavy-chain mRNA is observed in HeLa cells treated with TNF in combination with either type of IFN. To define the cis-acting elements mediating cytokine synergy, the promoter of a human major histocompatibility complex class I heavy-chain gene (HLA-B7) was placed in front of a reporter gene and transfected into HeLa cells. Deletion analysis mapped the elements required for synergy to a 40-bp region containing a kappa B-like element, which is necessary for the response to TNF, and an interferon consensus sequence (ICS), which is necessary for the responses to IFNs. When the orientation of these elements was reversed or their normal 20-bp spacing was reduced by 5 or 10 bp, i.e., one half or one full turn of the DNA helix, essentially equivalent responses were obtained, suggesting that these parameters are not critical. In electromobility shift assays, a p50-containing NF-kappa B nuclear factor from TNF-treated cells binds kappa B-containing probes, and ISGF-2 from IFN-gamma-treated cells binds ICS-containing probes. A probe containing both the kappa B and ICS elements (kappa B-ICS) forms a novel complex with nuclear factors isolated from cells treated with both TNF and IFN-gamma; this complex also forms when nuclear factors from individually cytokine-treated cells are mixed in vitro. The natural variant ICS found in HLA-A responds to IFN-gamma and can mediate synergy with TNF. However, the variant kappa B found in HLA-C does not respond to TNF, nor can it mediate synergy between TNF and IFN-gamma. These observations suggest that synergy between TNF and IFNs in the induction of HLA class I gene expression results from the sum of individual interactions of cytokine-activated enhancer-binding factors with the transcription initiation complex.
细胞因子肿瘤坏死因子（TNF）、β干扰素（IFN-β）和γ干扰素可增加主要组织相容性复合体I类分子的表达。在用TNF与任何一种干扰素联合处理的HeLa细胞中，观察到I类重链mRNA的诱导作用大于相加效应（即协同效应）。为了确定介导细胞因子协同作用的顺式作用元件，将人主要组织相容性复合体I类重链基因（HLA-B7）的启动子置于报告基因前，并转染到HeLa细胞中。缺失分析将协同作用所需的元件定位到一个40bp的区域，该区域包含一个对TNF应答所必需的κB样元件和一个对干扰素应答所必需的干扰素共有序列（ICS）。当这些元件的方向颠倒或其正常的20bp间距减少5或10bp，即DNA螺旋的半圈或一整圈时，获得了基本等效的应答，表明这些参数并不关键。在电泳迁移率变动分析中，来自TNF处理细胞的含p50的NF-κB核因子结合含κB的探针，来自γ干扰素处理细胞的ISGF-2结合含ICS的探针。一个同时包含κB和ICS元件（κB-ICS）的探针与从同时用TNF和γ干扰素处理的细胞中分离的核因子形成一种新型复合物；当将来自单独用细胞因子处理的细胞的核因子在体外混合时，也会形成这种复合物。在HLA-A中发现的天然变体ICS对γ干扰素作出应答，并可介导与TNF的协同作用。然而，在HLA-C中发现的变体κB对TNF无应答，也不能介导TNF与γ干扰素之间的协同作用。这些观察结果表明，TNF与干扰素在诱导HLA I类基因表达中的协同作用是细胞因子激活的增强子结合因子与转录起始复合物之间个体相互作用的总和。
