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首页> 外文期刊>Molecular and Cellular Biology >Four structurally distinct, non-DNA-binding subunits of human nuclear respiratory factor 2 share a conserved transcriptional activation domain.
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Four structurally distinct, non-DNA-binding subunits of human nuclear respiratory factor 2 share a conserved transcriptional activation domain.

机译：人类核呼吸因子2的四个结构上不同的非DNA结合亚基共享一个保守的转录激活域。

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Nuclear respiratory factor 2 (NRF-2) was previously purified to near homogeneity from HeLa cells on the basis of its ability to bind tandem recognition sites in the rat cytochrome oxidase subunit IV (RCO4) promoter. It consisted of five subunits, alpha, beta 1, beta 2, gamma 1, and gamma 2. Sequencing of tryptic peptides from alpha and from mixtures of the two beta or two gamma subunits revealed sequence identities with subunits of the mouse GA-binding protein (GABP), a ubiquitously expressed ETS domain activator composed of three subunits, alpha, beta 1, and beta 2. To understand the precise relationship between NRF-2 and GABP, cDNAs for all five NRF-2 subunits have now been cloned and their products have been overexpressed. The results establish that the two additional NRF-2 subunits are molecular variants that differ from GABP beta 1 and beta 2 by having a 12-amino-acid insertion containing two serine doublets. PCR and RNase protection assays show that mRNAs for these variants are expressed in the human but not the rodent cells and tissues examined. The insertion did not alter the ability of the beta and gamma subunits to associate with alpha, the DNA-binding subunit, nor did it affect the ability of NRF-2 beta 1 or beta 2 to direct high-affinity binding of alpha to tandem sites in the RCO4 promoter. In addition, the four NRF-2 beta and gamma subunits were equally proficient in activating transcription in transfected cells when fused to a GAL4 DNA-binding domain. The domain responsible for this transcriptional activation was localized by deletion mapping to a region of approximately 70 amino acids that is conserved in all four NRF-2 beta and gamma subunits. The repeated glutamine-containing hydrophobic clusters within this region bear a strong resemblance to those recently implicated in protein-protein interactions within the transcriptional apparatus.
机译：基于其结合大鼠细胞色素氧化酶亚基IV（RCO4）启动子中串联识别位点的能力，先前已从HeLa细胞中将核呼吸因子2（NRF-2）纯化至接近同质。它由五个亚基组成：α，β1，β2，γ1和γ2。从α以及两个β或两个γ亚基的混合物中对胰蛋白酶肽的测序揭示了与小鼠GA结合蛋白亚基的序列同一性。 （GABP），一种普遍表达的ETS域激活剂，由三个亚基（α，β1和β2组成）。要了解NRF-2和GABP之间的精确关系，现已克隆了所有五个NRF-2亚基的cDNA及其产品已被过分表达。结果证实，这两个另外的NRF-2亚基是分子变异体，与GABP beta 1和beta 2不同，它们具有包含两个丝氨酸双峰的12个氨基酸插入。 PCR和RNase保护试验表明，这些变体的mRNA在人类中表达，但在啮齿动物的细胞和组织中不表达。插入不会改变beta和γ亚基与DNA结合亚基alpha缔合的能力，也不会影响NRF-2 beta 1或beta 2指导alpha与串联位点高亲和力结合的能力。在RCO4启动子中。此外，当与GAL4 DNA结合结构域融合时，四个NRF-2β和γ亚基同样能有效激活转染细胞中的转录。通过缺失定位将负责该转录激活的结构域定位到在所有四个NRF-2β和γ亚基中保守的大约70个氨基酸的区域。在该区域内重复的含谷氨酰胺的疏水簇与最近与转录装置内的蛋白-蛋白相互作用有关的那些非常相似。

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