State Key Laboratory Breeding Base of Basic Science of Stomatology (Hubei-MOST) and Key Laboratory for Oral Biomedicine of Ministry of Education, School and Hospital of Stomatology, Wuhan University, Wuhan 430079, China.
J Biol Chem. 2013 Mar 29;288(13):9261-71. doi: 10.1074/jbc.M112.433730. Epub 2013 Feb 19.
Dentin tissue is derived from mesenchymal cells induced into the odontoblast lineage. The differentiation of odontoblasts is a complex process regulated by several transcriptional factor signaling transduction pathways. However, post-translational regulation of these factors during dentinogenesis remains unclear. To further explore the mechanisms, we investigated the role of microRNA (miRNA) during odontoblast differentiation. We profiled the miRNA expression pattern during mouse odontoblast differentiation using a microarray assay and identified that miR-145 and miR-143 were down-regulated during this process. In situ hybridization verified that the two miRNAs were gradually decreased during mouse odontoblast differentiation. Loss-of-function and gain-of-function experiments revealed that down-regulation of miR-145 and miR-143 could promote odontoblast differentiation and increased Dspp and Dmp1 expression in mouse primary dental pulp cells and vice versa. We found that miR-145 and miR-143 controlled odontoblast differentiation through several mechanisms. First, KLF4 and OSX bind to their motifs in Dspp and Dmp1 gene promoters and up-regulate their transcription thereby inducing odontoblast differentiation. The miR-145 binds to the 3'-UTRs of Klf4 and Osx genes, inhibiting their expression. Second, KLF4 repressed miR-143 transcription by binding to its motifs in miR-143 regulatory regions as detected by ChIP assay and dual luciferase reporter assay. Third, miR-143 regulates odontoblast differentiation in part through miR-145 pathway. Taken together, we for the first time showed that the miR-143 and miR-145 controlled odontoblast differentiation and dentin formation through KLF4 and OSX transcriptional factor signaling pathways.
牙本质组织来源于诱导牙胚谱系的间充质细胞。成牙本质细胞的分化是一个受多个转录因子信号转导通路调控的复杂过程。然而，牙本质发生过程中这些因子的翻译后调控仍不清楚。为了进一步探讨机制，我们研究了 miRNA（miRNA）在成牙本质细胞分化中的作用。我们通过微阵列分析对小鼠成牙本质细胞分化过程中的 miRNA 表达模式进行了分析，发现 miR-145 和 miR-143 在这个过程中下调。原位杂交验证了这两种 miRNA 在小鼠成牙本质细胞分化过程中逐渐减少。功能丧失和功能获得实验表明，下调 miR-145 和 miR-143 可促进小鼠原代牙髓细胞的成牙本质细胞分化，并增加 Dspp 和 Dmp1 的表达，反之亦然。我们发现 miR-145 和 miR-143 通过几种机制控制成牙本质细胞分化。首先，KLF4 和 OSX 结合到 Dspp 和 Dmp1 基因启动子中的其基序上，上调其转录，从而诱导成牙本质细胞分化。miR-145 结合到 Klf4 和 Osx 基因的 3'-UTR 上，抑制其表达。其次，ChIP 分析和双荧光素酶报告基因分析检测到，KLF4 通过结合 miR-143 调控区的基序抑制 miR-143 转录。第三，miR-143 通过 miR-145 通路部分调节成牙本质细胞分化。总之，我们首次表明 miR-143 和 miR-145 通过 KLF4 和 OSX 转录因子信号通路控制成牙本质细胞分化和牙本质形成。
