A family of C/EBP-related proteins capable of forming covalently linked leucine zipper dimers in vitro.
A family of C/EBP-related proteins capable of forming covalently linked leucine zipper dimers in vitro.
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DOI：
10.1101/gad.5.9.1553
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发表时间：
1991-09
影响因子：
10.5
通讯作者：
S. C. Williams;Carrie A. Cantwell;P. Johnson
中科院分区：
文献类型：
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作者：
S. C. Williams;Carrie A. Cantwell;P. Johnson
以c/ebp基因的DNA结合域为探针，通过降低严格性杂交筛选小鼠和大鼠基因组DNA文库。三个基因被分离出来，编码bZIP DNA结合蛋白（命名为CRP 1，CRP 2和CRP 3）具有很强的氨基酸序列相似性的C/EBP结合结构域。CRP 2与最近由其他组描述的蛋白质如NF-IL 6、IL 6、IL-6DBP和AGP/EBP相同，而CRP 1和CRP 3代表新的蛋白质。几条线索的证据表明，这三种蛋白质，沿着C/EBP，包括一个功能的家庭。每种细菌表达的多肽作为二聚体与DNA结合，其识别特性与C/EBP的识别特性几乎相同。每个成员还在羧基末端或其附近携带保守的半胱氨酸残基，紧接着亮氨酸拉链，其至少在体外允许配对拉链螺旋之间的有效二硫键交联。我们开发了一种共价二聚体的凝胶测定法，以评估家族成员之间的亮氨酸拉链特异性。结果表明，二聚体相互作用的所有成对组合都是可能的。在我们已经检查它们的程度上，在将适当的基因对共转染到受体细胞中后，可以在细胞内检测到相同的异二聚体复合物。所有成员也能够激活从含有C/EBP结合位点的启动子的体内转录。我们的研究结果表明，存在一组潜在的相互作用的C/EBP样蛋白，其复杂性与其他bZIP蛋白亚家族，如Jun，Fos和ATF/CREB。
Mouse and rat genomic DNA libraries were screened by reduced stringency hybridization with the DNA-binding domain of the c/ebp gene as a probe. Three genes were isolated that encode bZIP DNA-binding proteins (designated CRP1, CRP2, and CRP3) with strong amino acid sequence similarities to the C/EBP-binding domain. CRP2 is identical to the protein described recently by other groups as NF-IL6, LAP, IL-6DBP, and AGP/EBP, whereas CRP1 and CRP3 represent novel proteins. Several lines of evidence indicate that these three proteins, along with C/EBP, comprise a functional family. Each bacterially expressed polypeptide binds to DNA as a dimer with recognition properties that are virtually identical to those of C/EBP. Every member also bears a conserved cysteine residue at or near the carboxyl terminus, immediately following the leucine zipper, that at least in vitro allows efficient disulfide cross-linking between paired zipper helices. We developed a gel assay for covalent dimers to assess leucine zipper specificities among the family members. The results demonstrate that all pairwise combinations of dimer interactions are possible. To the extent that we have examined them, the same heterodimeric complexes can be detected intracellularly following cotransfection of the appropriate pair of genes into recipient cells. All members are also capable of activating in vivo transcription from promoters that contain a C/EBP-binding site. Our findings indicate that a set of potentially interacting C/EBP-like proteins exists, whose complexity is comparable to that of other bZIP protein subfamilies such as Jun, Fos, and ATF/CREB.