Protein and DNA contact surfaces that mediate the selective action of the Phox1 homeodomain at the c-fos serum response element
Protein and DNA contact surfaces that mediate the selective action of the Phox1 homeodomain at the c-fos serum response element
复制标题
DOI：
10.1128/mcb.17.11.6653
复制
发表时间：
1997-11-01
影响因子：
5.3
通讯作者：
Gilman, M
中科院分区：
文献类型：
--
作者：
Simon, KJ;Grueneberg, DA;Gilman, M
人类同源结构域蛋白Phox 1通过与血清应答因子（serum response factor，SRF）相互作用，赋予c-fos血清应答元件（serum response element，SRE）血清应答转录活性。这种活性与其他配对类同源结构域共享，但不与更远相关的同源结构域共享。为了理解Phox 1在SRE的作用机制以及在这种情况下配对类同源结构域的选择性活性的基础，我们对Phox 1同源结构域进行了详细的诱变。我们发现，氨基酸残基，接触的主要沟关闭的DNA所需的SRE激活在体内，这表明在体内需要的主要沟DNA接触的同源结构域。相比之下，Phox 1同源结构域的N-末端臂中的赖氨酸残基的取代似乎废除DNA结合而不影响体内活性。在螺旋1和2的暴露表面上的某些取代（DNA结合不需要）消除了体内活性，表明这些表面接触了该活性所需的辅助蛋白，我们还发现，从Phox 1螺旋1表面的单个氨基酸残基转移到远亲变形（Dfd）中的相应位置，同源结构域赋予Dfd在体内激活SRE的能力，我们认为Phox 1除了与SRF相互作用外，还与SRE上的一种或多种因子相互作用，并且这种相互作用的特异性由螺旋1和2表面上的残基决定。
The human homeodomain protein Phox1 can impart serum-responsive transcriptional activity to the c-fos serum response element (SRE) by interacting with serum response factor (SRF). This activity is shared with other Paired class homeodomains but not with more distantly related homeodomains, To understand the mechanism of action of Phox1 at the SRE and the basis for the selective activity of Paired class homeodomains in this context, we performed a detailed mutagenesis of the Phox1 homeodomain. We found that amino acid residues that contact the major groove off the DNA are required for SRE activation in vivo, suggesting an in vivo requirement for major-groove DNA contact by the homeodomain. In contrast, substitution of a lysine residue in the N-terminal arm of the Phox1 homeodomain appeared to abolish DNA binding without affecting activity in vivo. Certain substitutions on the exposed surfaces of helices 1 and 2, not required for DNA binding, abolished activity in vivo, suggesting that these surfaces contact an accessory protein(s) required for this activity, We also found that transfer of a single amino acid residue from the surface of Phox1 helix 1 to the corresponding position in the distantly related Deformed (Dfd) homeodomain imparts to Dfd the ability to activate the SRE in vivo, We propose that Phox1 interacts with one or more factors at the SRE, in addition to SRF, and that the specificity of this interaction is determined By residues on the surfaces of helices 1 and 2.