Abstract 1232: MicroRNA-1 Facilitates Myogenic Differentiation and Maturation into Skeletal Myotubes without Affecting Osteoblastic and Adipogenic Differentiation
MicroRNAs (miRNAs) are genomically encoded small noncoding RNAs, and emerging as an important and abundant class of post-transcriptional gene regulators. Recent evidences have shown that miRNAs are involved in the regulation of various cellular functions such as differentiation, cell growth and cell death, and a number of miRNAs exhibits tissue- or cell type-specific expression pattern. MiRNA-1 (miR-1) is a such tissue-specific miRNA, specifically expressed in hearts and skeletal muscles. In this study, we examined the role of miR-1 in myogenic, osteoblastic and adipogenic differentiation of C2C12 cells. Upon induction of myogenic differentiation, but not of osteoblastic differentiation, miR-1 was robustly expressed. To express exogenous mature miR-1 efficiently, we developed a retrovirus vector that expressed pre-miR-1 with additional 50-nt on each end under the control of RNA polymerase II promoter, forcing exogenous miRNA molecules to enter proper processing mechanisms. Retrovirus-mediated overexpression of miR-1 markedly enhanced myogenin expression in the early induction phase, which became comparable to that in mock-infected cells later. However, the expression of muscle creatine kinase, sarcomeric myosin and α-actinin was strikingly enhanced by miR-1 expression even in the late phase. Formation of myotubes was significantly augmented in miR-1-overexpressing cells, as shown by both the increased number of nuclei per myotube and the elevated ratio of myotube-incorporated nuclei to total nuclei. Thus, miR-1 expression enhanced not only differentiation but also maturation into mature myotubes. When osteoblastic differentiation was induced, the forced expression of miR-1 did not alter either expression of osteoblast markers such as osterix and osteocalcin, or ALP activity. Expression of adipocyte markers such as C/EBPα and PPARγ, and the lipid accumulation were also unaltered by miR-1 expression under adipogenic differentiation, indicating miR-1 did not influence the differentiation into these cell types. These results indicated that a muscle-specific miRNA, miR-1, plays an important role in controlling the myogenic differentiation and maturation in lineage-committed cells, rather than functioning in fate determination.