The Circular RNA circHUWE1 Sponges the miR-29b-AKT3 Axis to Regulate Myoblast Development

Abstract

Myogenesis is controlled by a well-established transcriptional hierarchy that coordinates the activities of a set of muscle genes. Recently, roles in myogenesis have been described for non-coding RNAs, including a role of circular RNA (circRNA) to regulate muscle gene expression. However, the functions of circRNA and the underlying mechanism by which circRNAs affect myogenesis remain poorly understood. In this study, we analyzed circRNA high-throughput sequencing results of bovine skeletal muscle samples and constructed a circRNA-miRNA-mRNA network according to the competitive endogenous RNA (ceRNA) theory. The putative circHUWE1-miR-29b-AKT3 network was analyzed and its involvement in myogenesis was confirmed through a series of assays. To assess the potential function of this regulation, bovine myoblasts were infected with overexpression plasmids and small interfering RNAs (siRNAs) that target circHUWE1. Next, cell proliferation, apoptosis, and differentiation were analyzed using Cell Counting Kit-8 (CCK-8), 5-ethynyl-2 0 -deoxyuridine (EdU), flow cytometry, western blotting, and qRT-PCR assays. The results suggest that circHUWE1 facilitates bovine myoblast proliferation and inhibits cell apoptosis and differentiation. Next, bioinformatics, dual-luciferase reporter assay, and AGO2 RNA immunoprecipitation (RIP) approaches were used to verify the interaction between circHUWE1, miR-29b, and AKT3. Subsequently, we identified that circHUWE1 could directly interfere with the ability of miR-29b to relieve AKT3 suppression, which ultimately activates the AKT signaling pathway. These findings suggested a new regulatory pathway for bovine skeletal muscle development, and they also expand our understanding of circRNA functions in mammals.

Circular RNA CircCOL5A1 Sponges the MiR-7-5p/Epac1 Axis to Promote the Progression of Keloids Through Regulating PI3K/Akt Signaling Pathway

“…For example, circCCDC9 acts as a ceRNA of miR-6792-3p to suppress the proliferation and invasion of gastric cancer cell lines in vitro and tumor growth and metastasis in vivo ( Luo et al, 2020 ). Yue et al (2020) constructed a circHUWE1-associated ceRNA network and then determined that circHUWE1 could directly sponge miR-29b to relieve AKT3 suppression, via activating the AKT signaling pathway in skeletal muscle development. In this research, we have studied a previously reported circRNA, named as circCOL5A1, which was derived from the host gene COL5A1 and was cyclized with the head-to-tail splicing of exon 13 and exon 19 ( Shi et al, 2020 ).…”

The Role of N6-Methyladenosine Modified Circular RNA in Pathophysiological Processes

“…Increasing evidences have shown that circRNAs are abundant in skeletal muscles during the differentiation myoblasts, and the global expression level of circRNA changes dynamically 153 . Additionally, several circRNAs (such as circ-ZNF609 and circular RNA supervillin have been proved played an important role in the differentiation and development of skeletal muscle 154 .…”

The Roles of CircRNAs in Regulating Muscle Development of Livestock Animals

“…Therefore, researchers have evaluated multiple co-regulatory relationships during muscle development. Among them, Yue et al (2020) verify the interaction among circHUWE1, miR-29b, and AKT3 with the help of bioinformatics, double luciferase report analysis, and AGO2-RNA immunoprecipitation (RIP) and point out that circHUWE1 can directly interfere with the ability of miR-29b to release AKT3 inhibition and finally activate the AKT signal pathway, thus promoting the proliferation of bovine myoblasts and inhibiting apoptosis and differentiation of bovine myoblasts. For this reason, Yue et al (2020) build upon previous research results to construct a ceRNA co-expression regulatory network, which not only provides a good idea for the study of muscle development, but also expands our understanding of the function of circRNAs.…”

“…Among them, Yue et al (2020) verify the interaction among circHUWE1, miR-29b, and AKT3 with the help of bioinformatics, double luciferase report analysis, and AGO2-RNA immunoprecipitation (RIP) and point out that circHUWE1 can directly interfere with the ability of miR-29b to release AKT3 inhibition and finally activate the AKT signal pathway, thus promoting the proliferation of bovine myoblasts and inhibiting apoptosis and differentiation of bovine myoblasts. For this reason, Yue et al (2020) build upon previous research results to construct a ceRNA co-expression regulatory network, which not only provides a good idea for the study of muscle development, but also expands our understanding of the function of circRNAs. KEGG pathway analysis shows that hosting genes of circRNAs are related to the muscle development pathway, including the mammalian target of rapamycin signaling pathway, Wnt signaling pathway, MAPK, and transforming growth factor-β signaling pathway ( Li et al, 2017 ).…”