title: miR-30 family members negatively regulate osteoblast differentiation.
abstract: miRNAs are endogenously expressed 18- to 25-nucleotide RNAs that regulate gene expression through translational repression by binding to a target mRNA. Recently, it has been indicated that miRNAs are closely related to osteogenesis. Our previous data suggested that miR-30 family members might be important regulators during the biomineralization process. However, whether and how they modulate osteogenic differentiation have not been explored. In this study, we demonstrated that miR-30 family members negatively regulate BMP-2-induced osteoblast differentiation by targeting Smad1 and Runx2. Evidentially, overexpression of miR-30 family members led to a decrease of alkaline phosphatase activity, whereas knockdown of them increased the activity. Then bioinformatic analysis identified potential target sites of the miR-30 family located in the 3' untranslated regions of Smad1 and Runx2. Western blot analysis and quantitative RT-PCR assays demonstrated that miR-30 family members inhibit Smad1 gene expression on the basis of repressing its translation. Furthermore, dual-luciferase reporter assays confirmed that Smad1 is a direct target of miR-30 family members. Rescue experiments that overexpress Smad1 and Runx2 significantly eliminated the inhibitory effect of miR-30 on osteogenic differentiation and provided strong evidence that miR-30 mediates the inhibition of osteogenesis by targeting Smad1 and Runx2. Also, the inhibitory effects of the miR-30 family were validated in mouse bone marrow mesenchymal stem cells. Therefore, our study uncovered that miR-30 family members are key negative regulators of BMP-2-mediated osteogenic differentiation.
authors.affiliation: Department of Prosthodontics, Shanghai Key Laboratory of Stomatology, 639 Zhi Zaoju Road, Shanghai 200011, China.
authors.forename: Tingting
authors.initials: T
authors.lastname: Wu
authors.forename: Haibo
authors.initials: H
authors.lastname: Zhou
authors.forename: Yongfeng
authors.initials: Y
authors.lastname: Hong
authors.forename: Jing
authors.initials: J
authors.lastname: Li
authors.forename: Xinquan
authors.initials: X
authors.lastname: Jiang
authors.forename: Hui
authors.initials: H
authors.lastname: Huang
mesh_terms: D000818:Animals; D055396:Bone Morphogenetic Protein 2 / Q000096:biosynthesis; D002454:Cell Differentiation / Q000502:physiology*; D002460:Cell Line; D050659:Core Binding Factor Alpha 1 Subunit / Q000096:biosynthesis; D005786:Gene Expression Regulation / Q000502:physiology*; D051379:Mice; D035683:MicroRNAs / Q000096:biosynthesis* / Q000235:genetics; D005810:Multigene Family / Q000502:physiology*; D010006:Osteoblasts / Q000166:cytology / Q000378:metabolism*; D014176:Protein Biosynthesis / Q000502:physiology*; D051898:Smad1 Protein / Q000096:biosynthesis
chemical_list: C525719:Bmp2 protein, mouse; D055396:Bone Morphogenetic Protein 2; D050659:Core Binding Factor Alpha 1 Subunit; D035683:MicroRNAs; C534715:Mirn30d microRNA, mouse; C493716:Runx2 protein, mouse; D051898:Smad1 Protein; C497401:Smad1 protein, mouse