Diamond Evan, Amen Melanie, Hu Qiaoyan, Espinoza Herbert M, Amendt Brad A
Center for Environmental and Genetic Medicine, Institute of Biosciences and Genetic Medicine Texas A&M Health Science Center, 2121 W. Holcombe Boulevard, Houston, TX 77030, USA.
Nucleic Acids Res. 2006;34(20):5951-65. doi: 10.1093/nar/gkl689. Epub 2006 Oct 26.
Dlx2, Lymphoid Enhancer Factor (Lef-1) and Msx2 transcription factors are required for several developmental processes. To understand the control of gene expression by these factors, chromatin immunoprecipitation (ChIP) assays identified Msx2 as a downstream target of Dlx2 and Lef-1. Dlx2 activates the Msx2 promoter in several cell lines and binds DNA as a monomer and dimer. A Lef-1 beta-catenin-dependent isoform minimally activates the Msx2 promoter and a Lef-1 beta-catenin-independent isoform is inactive, however co-expression of Dlx2 and both Lef-1 isoforms synergistically activate the Msx2 promoter. Co-immunoprecipitation and protein pull-down experiments demonstrate Lef-1 physically interacts with Dlx2. Deletion analyses of the Lef-1 protein reveal specific regions required for synergism with Dlx2. The Lef-1 beta-catenin binding domain (betaDB) is not required for its interaction with Dlx2. Msx2 can auto-regulate its promoter and repress Dlx2 activation. Msx2 repression of Dlx2 activation is dose-specific and both bind a common DNA-binding element. These transcriptional mechanisms correlate with the temporal and spatial expression of these factors and may provide a mechanism for the control of several developmental processes. We demonstrate new transcriptional activities for Dlx2, Msx2 and Lef-1 through protein interactions and identification of downstream targets.
Dlx2、淋巴样增强因子（Lef-1）和Msx2转录因子参与多个发育过程。为了解这些因子对基因表达的调控，染色质免疫沉淀（ChIP）分析确定Msx2是Dlx2和Lef-1的下游靶点。Dlx2在多种细胞系中激活Msx2启动子，并以单体和二聚体形式结合DNA。一种依赖β-连环蛋白的Lef-1亚型对Msx2启动子的激活作用最小，而一种不依赖β-连环蛋白的Lef-1亚型则无活性，然而，Dlx2与两种Lef-1亚型的共表达可协同激活Msx2启动子。免疫共沉淀和蛋白质下拉实验表明Lef-1与Dlx2存在物理相互作用。对Lef-1蛋白的缺失分析揭示了与Dlx2协同作用所需的特定区域。Lef-1的β-连环蛋白结合结构域（βDB）在其与Dlx2的相互作用中并非必需。Msx2可对其自身启动子进行自调控并抑制Dlx2的激活。Msx2对Dlx2激活的抑制具有剂量特异性，且二者均结合一个共同的DNA结合元件。这些转录机制与这些因子的时空表达相关，可能为多个发育过程的调控提供一种机制。我们通过蛋白质相互作用和下游靶点的鉴定，证明了Dlx2、Msx2和Lef-1具有新的转录活性。
