Kelm R J, Cogan J G, Elder P K, Strauch A R, Getz M J
Department of Biochemistry and Molecular Biology, Mayo Clinic/Foundation, Rochester, Minnesota 55905, USA.
J Biol Chem. 1999 May 14;274(20):14238-45. doi: 10.1074/jbc.274.20.14238.
Transcriptional activity of the mouse vascular smooth muscle alpha-actin gene in fibroblasts is regulated, in part, by a 30-base pair asymmetric polypurine-polypyrimidine tract containing an essential MCAT enhancer motif. The double-stranded form of this sequence serves as a binding site for a transcription enhancer factor 1-related protein while the separated single strands interact with two distinct DNA binding activities termed VACssBF1 and 2 (Cogan, J. G., Sun, S., Stoflet, E. S., Schmidt, L. J., Getz, M. J., and Strauch, A. R. (1995) J. Biol. Chem. 270, 11310-11321; Sun, S., Stoflet, E. S., Cogan, J. G., Strauch, A. R., and Getz, M. J. (1995) Mol. Cell. Biol. 15, 2429-2936). VACssBF2 has been recently cloned and shown to consist of two closely related proteins, Puralpha and Purbeta (Kelm, R. J., Elder, P. K., Strauch, A. R., and Getz, M. J. (1997) J. Biol. Chem. 272, 26727-26733). In this study, we demonstrate that Puralpha and Purbeta interact with each other via highly specific protein-protein interactions and bind to the purine-rich strand of the MCAT enhancer in the form of both homo- and heteromeric complexes. Moreover, both Pur proteins interact with MSY1, a VACssBF1-like protein cloned by virtue of its affinity for the pyrimidine-rich strand of the enhancer. Interactions between Puralpha, Purbeta, and MSY1 do not require the participation of DNA. Combinatorial interactions between these three single-stranded DNA-binding proteins may be important in regulating activity of the smooth muscle alpha-actin MCAT enhancer in fibroblasts.
小鼠血管平滑肌α-肌动蛋白基因在成纤维细胞中的转录活性部分受一个包含必需MCAT增强子基序的30个碱基对的不对称聚嘌呤-聚嘧啶序列调控。该序列的双链形式作为转录增强子因子1相关蛋白的结合位点，而分开的单链与两种不同的DNA结合活性相互作用，分别称为VACssBF1和2（科根，J.G.，孙，S.，斯托夫莱特，E.S.，施密特，L.J.，格茨，M.J.，和施特劳赫，A.R.（1995年）《生物化学杂志》270，11310 - 11321；孙，S.，斯托夫莱特，E.S.，科根，J.G.，施特劳赫，A.R.，和格茨，M.J.（1995年）《分子细胞生物学》15，2429 - 2936）。VACssBF2最近已被克隆，显示由两个密切相关的蛋白质Puralpha和Purbeta组成（凯尔姆，R.J.，埃尔德，P.K.，施特劳赫，A.R.，和格茨，M.J.（1997年）《生物化学杂志》272，26727 - 26733）。在本研究中，我们证明Puralpha和Purbeta通过高度特异性的蛋白质 - 蛋白质相互作用彼此相互作用，并以同聚和异聚复合物的形式结合到MCAT增强子的富含嘌呤的链上。此外，这两种Pur蛋白都与MSY1相互作用，MSY1是一种通过其对增强子富含嘧啶链的亲和力而克隆的VACssBF1样蛋白。Puralpha、Purbeta和MSY1之间的相互作用不需要DNA的参与。这三种单链DNA结合蛋白之间的组合相互作用可能在调节成纤维细胞中平滑肌α-肌动蛋白MCAT增强子的活性方面很重要。
