Bowlus C L, McQuillan J J, Dean D C
Department of Internal Medicine, Washington University School of Medicine, St. Louis, Missouri 63110.
J Biol Chem. 1991 Jan 15;266(2):1122-7.
A cAMP regulatory element (CRE) at nucleotide position -170 of the fibronectin gene was characterized previously (Dean, D. C., Blakeley, M. S., Newby, R. F., Ghazal, P., Hennighausen, L., and Bourgeois, S. (1989) Mol. Cell. Biol. 9, 1498-1506). Here we identify two additional low affinity CREs at nucleotide positions -260 and -415 which differ in sequence by 1 base pair. Interestingly, these CREs did not compete for binding of nuclear proteins in gel retardation assays and partial tryptic digestion of protein-DNA complexes produced a different pattern with each CRE, indicating that they bind different proteins. CRE (-170) competed for binding of proteins to both CREs, suggesting that it may represent a composite of the two elements. CRE (-415) competed effectively for binding of nuclear proteins to the somatostatin gene CRE, suggesting that, like the somatostatin CRE, it binds the nuclear protein CREB. On the other hand, CRE (-260) appears to bind the nuclear protein PEA-2, which also binds a site in the polyoma virus enhancer. In summary, disruption of dyad symmetry in the 3' region of the CRE, as occurs with CRE (-260) and CRE (-415), results in a lower affinity site and may also change the specificity for different nuclear proteins.
纤连蛋白基因核苷酸位置-170处的环磷酸腺苷（cAMP）反应元件（CRE）先前已被鉴定（迪恩，D.C.，布莱克利，M.S.，纽比，R.F.，加扎尔，P.，亨尼豪森，L.，和布尔乔亚，S.（1989年）《分子与细胞生物学》9，1498 - 1506）。在此，我们在核苷酸位置-260和-415处鉴定出另外两个低亲和力CRE，它们在序列上相差1个碱基对。有趣的是，在凝胶阻滞分析中，这些CRE不竞争核蛋白的结合，并且蛋白质-DNA复合物的部分胰蛋白酶消化对每个CRE产生了不同的模式，表明它们结合不同的蛋白质。CRE（-170）竞争蛋白质与两个CRE的结合，这表明它可能代表这两个元件的组合。CRE（-415）有效地竞争核蛋白与生长抑素基因CRE的结合，这表明，与生长抑素CRE一样，它结合核蛋白CREB。另一方面，CRE（-260）似乎结合核蛋白PEA - 2，PEA - 2也结合多瘤病毒增强子中的一个位点。总之，如CRE（-260）和CRE（-415）那样，CRE 3'区域中二联体对称性的破坏会导致一个低亲和力位点，并且还可能改变对不同核蛋白的特异性。
