title: MicroRNA-590-5p Stabilizes Runx2 by Targeting Smad7 During Osteoblast Differentiation.
abstract: Mesenchymal stem cells (MSCs) are multipotent cells and their differentiation into the osteoblastic lineage is strictly controlled by several regulators, including microRNAs (miRNAs). Runx2 is a bone transcription factor required for osteoblast differentiation. Here, we used in silico analysis to identify a number of miRNAs that putatively target Runx2 and its co-factors to mediate both positive and negative regulation of osteoblast differentiation. Among these miRNAs, miR-590-5p was selected and its expression was found to be increased during osteoblast differentiation. When mouse MSCs (mMSCs) were transiently transfected with a miR-590-5p mimic, we detected an increase in both calcium deposition and the mRNA expression of osteoblast differentiation marker genes such as alkaline phosphatase (ALP) and type I collagen genes. Smad7 was found to be among the putative target genes of miR-590-5p and its mRNA and protein expression decreased after miR-590-5p mimic transfection in human osteoblast-like cells (MG63). Our analysis indicated that Runx2 was not a putative target of miR-590-5p. However, Runx2 protein, but not mRNA expression, increased after miR-590-5p mimic transfection in MG63 cells. Runx2 protein expression was increased with knockdown of Smad7 expression by Smad7 siRNA in these cells. We further identified that the 3'-untranslated region of Smad7 was directly targeted by miR-590-5p; this was done using the luciferase reporter gene system. It is known that Smad7 inhibits osteoblast differentiation via Smurf2-mediated Runx2 degradation. Hence, based on our results, we suggest that miR-590-5p promotes osteoblast differentiation by indirectly protecting and stabilizing the Runx2 protein by targeting Smad7 gene expression. J. Cell. Physiol. 232: 371-380, 2017. © 2016 Wiley Periodicals, Inc.
authors.affiliation: Department of Biotechnology, School of Bioengineering, SRM University, Kattankulathur, Tamil Nadu, India.
authors.forename: M
authors.initials: M
authors.lastname: Vishal
authors.affiliation: Department of Biotechnology, School of Bioengineering, SRM University, Kattankulathur, Tamil Nadu, India.
authors.forename: S
authors.initials: S
authors.lastname: Vimalraj
authors.affiliation: Department of Biotechnology, School of Bioengineering, SRM University, Kattankulathur, Tamil Nadu, India.
authors.forename: R
authors.initials: R
authors.lastname: Ajeetha
authors.affiliation: Department of Biotechnology, School of Bioengineering, SRM University, Kattankulathur, Tamil Nadu, India.
authors.forename: M
authors.initials: M
authors.lastname: Gokulnath
authors.affiliation: Department of Biotechnology, School of Bioengineering, SRM University, Kattankulathur, Tamil Nadu, India.
authors.forename: R
authors.initials: R
authors.lastname: Keerthana
authors.affiliation: Department of Basic and Craniofacial Biology, New York University College of Dentistry, New York, New York.
authors.forename: Z
authors.initials: Z
authors.lastname: He
authors.affiliation: Department of Basic and Craniofacial Biology, New York University College of Dentistry, New York, New York.
authors.forename: N C
authors.initials: NC
authors.lastname: Partridge
authors.affiliation: Department of Biotechnology, School of Bioengineering, SRM University, Kattankulathur, Tamil Nadu, India. selvamurugan.n@ktr.srmuniv.ac.in.
authors.forename: N
authors.initials: N
authors.lastname: Selvamurugan
mesh_terms: D000328:Adult; D001483:Base Sequence; D002454:Cell Differentiation / Q000235:genetics*; D003198:Computer Simulation; D050659:Core Binding Factor Alpha 1 Subunit / Q000235:genetics / Q000378:metabolism*; D015536:Down-Regulation / Q000235:genetics; D005260:Female; D006801:Humans; D035683:MicroRNAs / Q000378:metabolism*; D008954:Models, Biological; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism*; D055550:Protein Stability; D051906:Smad7 Protein / Q000235:genetics / Q000378:metabolism*
chemical_list: D050659:Core Binding Factor Alpha 1 Subunit; C568221:MIRN590 microRNA, human; D035683:MicroRNAs; D051906:Smad7 Protein