de Crombrugghe B, Lefebvre V, Behringer R R, Bi W, Murakami S, Huang W
Department of Molecular Genetics, The University of Texas, M.D. Anderson Cancer Center, 1515 Holcombe Blvd., Houston, TX 77030, USA.
Matrix Biol. 2000 Sep;19(5):389-94. doi: 10.1016/s0945-053x(00)00094-9.
With the goal of identifying master transcription factors that control the genetic program of differentiation of mesenchymal cells into chondrocytes, we first delineated a 48-bp chondrocyte-specific enhancer element in the gene for proalpha1(II) collagen (Col2a1), an early and abundant marker of chondrocytes. Our experiments have demonstrated that the HMG-box-containing transcription factor, Sox9 which binds and activates this enhancer element, is required for chondrocyte differentiation and for expression of a series of chondrocyte-specific marker genes including Col2a1, Col9a2, Col11a2 and Aggrecan. In the absence of Sox9 the block in differentiation occurs at the stage of mesenchymal condensation, suggesting the hypothesis that Sox9 might also control expression of cell surface proteins needed for mesenchymal condensation. Since Sox9 also contains a potent transcription activation domain, it is a typical transcription factor. Two other members of the Sox family, L-Sox5 and Sox6, also bind to the 48-bp Col2a1 enhancer and together with Sox9 activate this enhancer as well as the endogenous Col2a1 and aggrecan genes. L-Sox5 and Sox6 have a high degree of sequence identity to each other and are likely to have redundant functions. Except for the HMG-box, L-Sox5 and Sox6 have no similarity to Sox9 and, hence, are likely to have a complementary function to that of Sox9. Our experiments suggest the hypothesis that, like Sox9, Sox5 and Sox6 might also be needed for chondrocyte differentiation. Other experiments, have provided evidence that the Sox9 polypeptide and the Sox9 gene are targets of signaling molecules that are known to control discrete steps of chondrogenesis in the growth plate of endochondral bones. Protein kinase A (PKA) phosphorylation of Sox9 increases its DNA binding and transcriptional activity. Since PKA-phosphorylated-Sox9 is found in the prehypertrophic zone of the growth plate, the same location where the gene for the receptor of the parathyroid hormone-related peptide (PTHrP) is expressed and since PTHrP signaling is mediated by cyclic AMP, we have hypothesized that Sox9 is a target for PTHrP signaling. Other experiments have also shown that fibroblast growth factors (FGFs) increase the expression of Sox9 in chondrocytes in culture and that this activation is mediated by the mitogen-activated protein kinase pathway. These results favor the hypothesis that in achondroplasia, a disease caused by activating mutations in FGF receptor 3, there might also be an abnormally high Sox9 expression.
为了确定控制间充质细胞分化为软骨细胞这一遗传程序的主要转录因子，我们首先在编码原α1（II）型胶原蛋白（Col2a1）的基因中划定了一个48bp的软骨细胞特异性增强子元件，Col2a1是软骨细胞早期且丰富表达的标志物。我们的实验表明，含HMG盒的转录因子Sox9可结合并激活该增强子元件，它是软骨细胞分化以及包括Col2a1、Col9a2、Col11a2和聚集蛋白聚糖在内的一系列软骨细胞特异性标志物基因表达所必需的。在没有Sox9的情况下，分化阻滞发生在间充质凝聚阶段，这提示了一个假说，即Sox9可能还控制间充质凝聚所需的细胞表面蛋白的表达。由于Sox9还含有一个有效的转录激活结构域，它是一个典型的转录因子。Sox家族的另外两个成员L-Sox5和Sox6也与48bp的Col2a1增强子结合，并与Sox9一起激活该增强子以及内源性的Col2a1和聚集蛋白聚糖基因。L-Sox5和Sox6彼此具有高度的序列同一性，可能具有冗余功能。除了HMG盒外，L-Sox5和Sox6与Sox9没有相似性，因此，它们可能具有与Sox9互补的功能。我们的实验提示了一个假说，即与Sox9一样，Sox5和Sox6可能也是软骨细胞分化所必需的。其他实验提供了证据表明，Sox9多肽和Sox9基因是已知控制软骨内骨生长板软骨形成离散步骤的信号分子的靶点。蛋白激酶A（PKA）对Sox9的磷酸化增加了其DNA结合和转录活性。由于PKA磷酸化的Sox9存在于生长板的前肥大区，甲状旁腺激素相关肽（PTHrP）受体基因也在同一位置表达，并且由于PTHrP信号是由环磷酸腺苷介导的，我们推测Sox9是PTHrP信号的靶点。其他实验还表明，成纤维细胞生长因子（FGFs）可增加培养的软骨细胞中Sox9的表达，并且这种激活是由丝裂原活化蛋白激酶途径介导的。这些结果支持了这样一个假说，即在由FGF受体3激活突变引起的疾病软骨发育不全中，也可能存在异常高的Sox9表达。
