Title

The Role of microRNAs in Bone

Date of Completion

January 2010

Keywords

Biology, Molecular|Biology, Cell

Degree

Ph.D.

Abstract

miRNAs are short, non-coding RNAs that negatively regulate gene expression by binding to specific sequences within a target mRNA. Families of miRNAs can work coorperatively to regulate complex cellular processes, including differentiation (Chapter 1). The purpose of these studies was to understand the molecular mechanisms by which miRNAs regulate osteoblast differentiation. ^ miR-29 Suppression of Osteonectin in Osteoblasts: Regulation During Differentiation and by Canonical Wnt Signaling (Chapter 3). Osteonectin is expressed by osteoblasts and regulates collagen fibril formation. In osteoblasts, osteonectin expression during matrix deposition decreases with matrix maturation/mineralization. Osteonectin is a target for miR-29, and miR-29 levels are low during matrix deposition, and increase as osteoblasts mature. Canonical Wnt signaling increases as osteoblasts mature, providing a potential mechanism for downregulating osteonectin during differentiation. Further, Wnt signaling rapidly induces miR-29 and decreases osteonectin. ^ Single nucleotide polymorphisms in the human osteonectin/SPARC 3' UTR mediate differential regulation of gene expression: regulation by microRNAs (Chapter 4). Previously, haplotypes consisting of 3 SNPs in the 3'UTR of osteonectin, at cDNA bases 1046(C/G), 1599(C/G) and 1970(T/G), are found to be associated with bone mass in a cohort of men with idiopathic osteoporosis. Interestingly, the osteonectin 3' UTR haplotype found at the highest frequency in the most severely affected osteoporosis patients allowed the lowest level of gene expression. We demonstrated that SNPs at 1599 and 1970 differentially regulate gene expression, and characterized miRNAs that may selectively interact with SNP regions. Selective regulation of osteonectin by miRNAs could be one mechanism contributing to variation in bone mass. ^ miR-29a modulates canonical Wnt signaling in human osteoblasts through a positive feedback loop (Chapter 5). We found that miR-29a transcription is induced by canonical Wnt signaling. miR-29a promotes the expression of osteoblastic markers, and directly targets Dkk1, Kremen2, and sFRP2, inhibitors of Wnt signaling. This results in miR-29a potentiating Wnt signaling, to drive osteoblast differentiation.^ The study of miRNAs in bone cells is novel and miRNA-based therapeutics may be promising for the treatment of bone diseases (Chapter 6). ^