Santoro I M, Walsh K
Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio 44106.
Mol Cell Biol. 1991 Dec;11(12):6296-305. doi: 10.1128/mcb.11.12.6296-6305.1991.
DNA elements with the CC(A/T)6GG, or CArG, motif occur in promoters that are under different regulatory controls. CArG elements from the skeletal actin, c-fos, and myogenin genes were tested for their abilities to confer tissue-specific expression on reporter genes when the individual elements were situated immediately upstream from a TATA element. The c-fos CArG element, also referred to as the serum response element (SRE), conferred basal, constitutive expression on the test promoter. The CArG motif from the myogenin gene was inactive. The skeletal actin CArG motif functioned as a muscle regulatory element (MRE) in that basal expression was detected only in muscle cultures. Muscle-specific expression from the 28-bp MRE and the 2.3-kb skeletal actin promoter was trans repressed by the Fos and Jun proteins. The expression and factor-binding properties of a series of synthetic CArG elements were analyzed. Muscle-specific expression was conferred by perfect 28-bp palindromes on the left and right halves of the skeletal actin MRE. Chimeric elements of the skeletal actin MRE and the c-fos SRE differed in their expression properties. Muscle-specific expression was observed when the left half of the MRE was fused to the right half of the SRE. Constitutive expression was conferred by a chimera with the right half of the MRE fused to the left half of the SRE and by chimeras which exchanged the central CC(A/T)6GG sequences. At least three distinct proteins specifically bound to these CArG elements. The natural and synthetic CArG elements differed in their affinities for these proteins; however, muscle-specific expression could not be attributed to differences in the binding of a single protein. Furthermore, the MRE did not bind MyoD or the myogenin-E12 heterodimer, indicating that muscle-specific expression from this element does not involve a direct interaction with these helix-loop-helix proteins. These data demonstrate that the conserved CArG motifs form the core of a family of functionally different DNA regulatory elements that may contribute to the tissue-specific expression properties of their cognate promoters.
具有CC(A/T)6GG基序（即CArG基序）的DNA元件存在于受不同调控的启动子中。当单个元件位于TATA元件上游紧邻位置时，对来自骨骼肌肌动蛋白、c-fos和肌细胞生成素基因的CArG元件赋予报告基因组织特异性表达的能力进行了测试。c-fos CArG元件，也称为血清反应元件（SRE），赋予测试启动子基础的、组成型表达。来自肌细胞生成素基因的CArG基序无活性。骨骼肌肌动蛋白CArG基序作为肌肉调节元件（MRE）发挥作用，因为仅在肌肉培养物中检测到基础表达。28个碱基对的MRE和2.3千碱基对的骨骼肌肌动蛋白启动子的肌肉特异性表达受到Fos和Jun蛋白的反式抑制。分析了一系列合成CArG元件的表达和因子结合特性。骨骼肌肌动蛋白MRE左右两半的完美28个碱基对回文结构赋予肌肉特异性表达。骨骼肌肌动蛋白MRE和c-fos SRE的嵌合元件在表达特性上有所不同。当MRE的左半部分与SRE的右半部分融合时，观察到肌肉特异性表达。由MRE的右半部分与SRE的左半部分融合而成的嵌合体以及交换了中央CC(A/T)6GG序列的嵌合体赋予组成型表达。至少有三种不同的蛋白质特异性结合这些CArG元件。天然和合成的CArG元件对这些蛋白质的亲和力不同；然而，肌肉特异性表达不能归因于单一蛋白质结合的差异。此外，MRE不结合MyoD或肌细胞生成素-E12异二聚体，表明该元件的肌肉特异性表达不涉及与这些螺旋-环-螺旋蛋白的直接相互作用。这些数据表明，保守的CArG基序构成了一个功能不同的DNA调节元件家族的核心，这些元件可能有助于其同源启动子的组织特异性表达特性。
