Regulation of Tcf7 by Runx2 in chondrocyte maturation and proliferation
Regulation of Tcf7 by Runx2 in chondrocyte maturation and proliferation
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DOI：
10.1007/s00774-010-0222-z
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发表时间：
2011-05-01
影响因子：
3.3
通讯作者：
Komori, Toshihisa
中科院分区：
文献类型：
--
作者：
Mikasa, Masaki;Rokutanda, Satoshi;Komori, Toshihisa
Runx 2在软骨细胞分化和增殖的调控中起重要作用，但Runx 2的靶分子仍有待研究。我们使用微阵列搜索通过将Runx 2引入Runx 2（-/-）软骨细胞而上调的基因，发现Tcf 7被Runx 2上调。因此，我们研究了Runx 2在调节Tcf/Lef家族转录因子中的功能。Runx 2强烈诱导Tcf 7和Lef 1，但Tcf 7 l1和Tcf 7 l2在Runx 2（-/-）软骨细胞中仅轻微诱导; Tcf 7和Tcf 7 l2在Runx 2（-/-）软骨骨骼和颅骨中的表达降低，并且Tcf 7表现出与Runx 2相似的表达模式。在报告基因测定中，Runx 2温和地激活了8.6和1.8 kb的Tcf 7启动子构建体。使用1.8 kb片段的缺失构建体的报告基因测定表明，0.3 kb启动子区负责Runx 2依赖性转录激活。为了研究Tcf 7在骨骼发育中的功能，我们使用Col 2a 1启动子产生显性阴性（dn）Tcf 7转基因小鼠。Dn-Tcf 7转基因胚胎表现出侏儒症，并且在四肢、肋骨和椎骨中以依赖于转基因表达水平的方式延迟矿化。原位杂交分析表明，软骨内骨化延迟dn-Tcf 7转基因胚胎由于减速软骨细胞成熟。此外，BrdU标记显示dn-Tcf 7转基因胚胎中生长板增殖层中软骨细胞增殖减少。这些发现表明Runx 2至少部分地通过Tcf 7的诱导来调节软骨细胞的成熟和增殖。
Runx2 plays important roles in the regulation of chondrocyte differentiation and proliferation; however, the Runx2 target molecules still remain to be investigated. We searched the genes upregulated by the introduction of Runx2 into Runx2 (-/-) chondrocytes using microarray and found that Tcf7 is upregulated by Runx2. Thus, we examined the functions of Runx2 in the regulation of the Tcf/Lef family of transcription factors. Runx2 induced Tcf7 and Lef1 strongly, but Tcf7l1 and Tcf7l2 only slightly in Runx2 (-/-) chondrocytes; the expressions of Tcf7 and Tcf7l2 were reduced in Runx2 (-/-) cartilaginous skeletons and calvaria, and Tcf7 showed a similar expression pattern to Runx2. In reporter assays, Runx2 mildly activated the 8.6 and 1.8 kb Tcf7 promoter constructs. The reporter assays using the deletion constructs of the 1.8-kb fragment showed that the 0.3-kb promoter region is responsible for the Runx2-dependent transcriptional activation. To investigate the function of Tcf7 in skeletal development, we generated dominant-negative (dn) Tcf7 transgenic mice using the Col2a1 promoter. Dn-Tcf7 transgenic embryos showed dwarfism, and mineralization was retarded in limbs, ribs, and vertebrae in a manner dependent on the expression levels of the transgene. In situ hybridization analysis showed that endochondral ossification is retarded in dn-Tcf7 transgenic embryos due to the decelerated chondrocyte maturation. Further, BrdU labeling showed a reduction in chondrocyte proliferation in the proliferating layer of the growth plate in dn-Tcf7 transgenic embryos. These findings indicate that Runx2 regulates chondrocyte maturation and proliferation at least partly through the induction of Tcf7.