05 April 2019 : Laboratory Research
Hsa_circ_0002468 Regulates the Neuronal Differentiation of SH-SY5Y Cells by Modulating the MiR-561/E2F8 Axis
Minhui Yang1ABCDEFG, Guanghong Xiang2BE, Dan Yu3CF, Guoshuai Yang3CDF, Weifeng He4BF, Songlin Yang2DF, Gaoya Zhou2ABE, Aiqun Liu5ABCDEFG*DOI: 10.12659/MSM.915518
Med Sci Monit 2019; 25:2511-2519
Abstract
BACKGROUND: It has been shown that circular RNAs (circRNAs) play a vital role in the regulation of neuronal differentiation; however, the precise role of circRNAs in human neuronal differentiation remains largely unexplored.
MATERIAL AND METHODS: A dual-luciferase reporter assay was carried out to confirm the targets of hsa_circ_0002468, miR-561, E2F8 (E2F transcription factor 8, a protein coding gene), and miR-561. We detected the expression of hsa_circ_0002468, miR-561, and E2F8 by using quantitative real-time polymerase chain reaction (qRT-PCR) analyses. In addition, we performed the functional experiments by using a BrdU (5-bromo-2’-deoxyuridine) assay and qRT-PCR analyses.
RESULTS: In this study, we showed that hsa_circ_0002468 can act as a sponge of miR-561 to regulate SH-SY5Y proliferation and differentiation. A bioinformatics analysis showed that hsa_circ_0002468 had a binding site that corresponded to miR-561, which was verified by dual-luciferase reporter assay. The expression of hsa_circ_0002468 was increased during SH-SY5Y differentiation and was inversely correlated with miR-561 expression. Using qRT-PCR analysis, we showed that hsa_circ_0002468 negatively regulated miR-561 in SH-SY5Y cells. Intriguingly, the overexpression of hsa_circ_0002468 increased SH-SY5Y differentiation and reduced SH-SY5Y proliferation; the suppression of hsa_circ_0002468 led to decreased SH-SY5Y differentiation levels and increased SH-SY5Y proliferation levels. Additionally, overexpression of miR-561 rescued the SH-SY5Y proliferation deficiency induced by hsa_circ_0002468 overexpression and abolished the SH-SY5Y differentiation promoted by hsa_circ_0002468. Furthermore, E2F8 was validated as a direct target of miR-561.
CONCLUSIONS: Our data suggested that hsa_circ_0002468 was a novel circRNA that regulated SH-SY5Y cell proliferation and differentiation via targeting the miR-561/E2F8 axis. Therefore, manipulating hsa_circ_0002468 in SH-SY5Y cells could be a novel strategy to develop novel interventions for the treatment of relevant neurological disorders.
Keywords: E2F6 Transcription Factor, Neuregulins, RNA, Untranslated, Cell Line, Computational Biology, Neurons, RNA, RNA, Circular, Real-Time Polymerase Chain Reaction, Repressor Proteins
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