MicroRNAs (miRNAs) have been found to play essential roles in muscle cell proliferation and differentiation. MicroRNA-1 (miR-1) and microRNA-206 (miR-206), which are similar and have the same seed sequence, have specific roles in modulating skeletal muscle proliferation and differentiation in vitro and in vivo. However, there is no information about their function during bovine skeletal muscle satellite cell development. In this study, the profiles of miR-1 and miR-206 and their biological functions in bovine skeletal muscle cell development was investigated. The target genes were predicted, and we used a dual-luciferase reporter assay to demonstrate that miR-1 and miR-206 directly targeted the 3' untranslated region (3'UTR) of paired-box transcription factor Pax7 and histone deacetylase 4 (HDAC4). We showed that miR-1 and miR-206 facilitate bovine skeletal muscle satellite cell myogenic differentiation by restricting the expression of their target gene and that inhibition of miR-1 and miR-206 increased the Pax7 and HDAC4 protein levels and substantially enhanced satellite cell proliferation. Therefore, our results revealed the mechanism in which miR-1 and miR-206 positively regulate bovine skeletal muscle satellite cell myogenic differentiation via Pax7 and HDAC4 downregulation.

MicroRNAs (miRNAs) play essential roles in muscle cell proliferation and differentiation. The muscle-specific miRNAs miR-1 and miR-206 have been shown to regulate muscle development and promote myogenic differentiation; however, it is likely that a number of other miRNAs play important roles in regulating myogenesis as well. microRNA-128 (miR-128) has been reported to be highly expressed in brain and skeletal muscle, and we found that miR-128 is also up-regulated during bovine skeletal muscle satellite cell differentiation using microarray analysis and qRT-PCR. However, little is known about the functions of miR-128 in bovine skeletal muscle satellite cell development. In this study, we investigated the biological functions of miR-128 in bovine skeletal muscle cell development. Using a dual-luciferase reporter assay, we confirmed that miR-128 regulates the Sp1 gene. Over-expression of miR-128 reduced Sp1 protein levels and inhibited muscle satellite cell proliferation and differentiation. Inhibition of miR-128 increased Sp1 protein levels and promoted muscle satellite cell differentiation but also suppressed proliferation. Changes in miR-128 and Sp1 expression levels also affected the protein levels of MyoD and CDKN1A. Sp1, an activator of MyoD and a suppressor of CDKN1A, plays an important role in bovine muscle cell proliferation and differentiation. The results of our study reveal a mechanism by which miR-128 regulates bovine skeletal muscle satellite cell proliferation and myogenic differentiation via Sp1.

Development of skeletal muscle is a complicated biological process regulated by various regulation factors and signal pathways. MicroRNAs (miRNAs) are novel gene regulators that control muscle cell development. microRNA-143 (miR-143) is highly expressed in skeletal muscle, and we found that miR-143 level is significantly increased during bovine skeletal muscle satellite cells (MSCs) differentiation process through microarray analysis and qRT-PCR detection. However, the function of miR-143 in bovine muscle development remained unclear. In our work, the functions of miR-143 in bovine MSCs myogenic differentiation were investigated. We discovered that IGFBP5 is directly regulated by miR-143 using a dual-luciferase reporter assay. Overexpression of miR-143 led to decreased level of IGFBP5 protein and restrained cell proliferation and differentiation, while downregulation of miR-143 resulted in increased levels of IGFBP5 protein and restrained cell proliferation but improved differentiation. IGFBP5, an important component of IGF signaling pathway, contributes greatly to bovine muscle cell development. A mechanism that miR-143 can regulate the proliferation and differentiation of bovine MSCs through changing expression of IGFBP5 was elucidated by our study.

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