title: Effects of miR-335-5p in modulating osteogenic differentiation by specifically downregulating Wnt antagonist DKK1.
abstract: Dickkopf-related protein 1 (DKK1) is essential to maintain skeletal homeostasis as an inhibitor of Wnt signaling and osteogenic differentiation. The purpose of this study was to investigate the molecular mechanisms underlying the developmental stage-specific regulation of the DKK1 protein level. We performed a series of studies including luciferase reporter assays, micro-RNA microarray, site-specific mutations, and gain- and loss-of-function analyses. We found that the DKK1 protein level was regulated via DKK1 3' UTR by miRNA control, which was restricted to osteoblast-lineage cells. As a result of decreased DKK1 protein level by miR-335-5p, Wnt signaling was enhanced, as indicated by elevated GSK-3β phosphorylation and increased β-catenin transcriptional activity. The effects of miR-335-5p were reversed by anti-miR-335-5p treatment, which downregulated endogenous miR-335-5p. In vivo studies showed high expression levels of miR-335-5p in osteoblasts and hypertrophic chondrocytes of mouse embryos, indicating a pivotal role of miR-335-5p in regulating bone development. In conclusion, miR-335-5p activates Wnt signaling and promotes osteogenic differentiation by downregulating DKK1. This cell- and development-specific regulation is essential and mandatory for the initiation and progression of osteogenic differentiation. miR-335-5p proves to be a potential and useful targeting molecule for promoting bone formation and regeneration.
authors.affiliation: Division of Oral Biology, Tufts University School of Dental Medicine, Boston, MA 02111, USA.
authors.forename: Jin
authors.initials: J
authors.lastname: Zhang
authors.forename: Qisheng
authors.initials: Q
authors.lastname: Tu
authors.forename: Lynda F
authors.initials: LF
authors.lastname: Bonewald
authors.forename: Xi
authors.initials: X
authors.lastname: He
authors.forename: Gary
authors.initials: G
authors.lastname: Stein
authors.forename: Jane
authors.initials: J
authors.lastname: Lian
authors.forename: Jake
authors.initials: J
authors.lastname: Chen
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics; D000818:Animals; D002454:Cell Differentiation / Q000235:genetics*; D002460:Cell Line; D015536:Down-Regulation / Q000235:genetics*; D004622:Embryo, Mammalian / Q000166:cytology / Q000378:metabolism; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism*; D051379:Mice; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis; D010006:Osteoblasts / Q000378:metabolism; D010012:Osteogenesis / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000235:genetics; D051153:Wnt Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: D020413:3' Untranslated Regions; C501127:Dkk1 protein, mouse; D036341:Intercellular Signaling Peptides and Proteins; D035683:MicroRNAs; C541986:Mirn335 microRNA, mouse; D051153:Wnt Proteins