Abstract
Neural stem cells (NSCs) serve as the source of both neurons and support cells, and neurogenesis is reportedly linked to the circadian clock. This study aimed to clarify the functional role of the circadian rhythm-related nuclear receptor, REV-ERBβ, in neurogenesis of NSCs from adult brain. Accordingly, Rev-erbβ expression and the effect of Rev-erbβ gene-specific knockdown on neurogenesis in vitro was examined in adult rodent NSCs. Initial experiments confirmed REV-ERBβ expression in cultured adult NSCs, while subsequent gene expression and gene ontogeny analyses identified functional genes upregulated or downregulated by REV-ERBβ. In particular, expression levels of factors associated with proliferation, stemness, and neural differentiation were affected. Knockdown of Rev-erbβ showed involvement of REV-ERBβ in regulation of cellular proliferation and self-renewal of cultured adult NSCs. Moreover, Rev-erbβ-knockdown cells formed neurons with a slightly shrunken morphology, fewer new primary neurites, and reduced length and branch formation of neurites. Altogether, this suggests that REV-ERBβ is involved in neurite formation during neuronal differentiation of cultured adult NSCs. In summary, REV-ERBβ is a known circadian regulatory protein that appears to be involved in neurogenesis via regulation of networks for cell proliferation and neural differentiation/maturation in adult NSCs.
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12 March 2021
A Correction to this paper has been published: https://doi.org/10.1007/s10571-021-01076-5
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Acknowledgements
I thank K. Akagi and the members of the Life Science Support Center at Nagasaki University for technical assistance and helpful discussions. Editage by Cactus and Edanz Group Japan are acknowledged for their assistance with preparation of the manuscript. I also thank Alison Sherwin, Ph.D., from Edanz Group (www.edanzediting.com/ac) for editing a draft of the manuscript. This work was supported by a Grant-in-Aid for Scientific Research (C) (Grant Number 15K06711) from the Japan Society for the Promotion of Science (JSPS) and a Grant-in-Aid from the Alumni Association of Nagasaki University School of Medicine.
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The original online version of this article was revised: the keyword "REV-ERBβ" has been corrected as "REV-ERB".
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Supplementary material 1 (PDF 1184 kb) Supplementary Material 1 Quantification of neural differentiation efficiency of mouse and rat cultured adult neural stem cells (NSCs) after gene-specific knockdown. Percentage of GFP and highly expressed GFAP (GFAPhigh) or βIII-tubulin double-positive cells in mouse (a) or rat (b) adult NSCs after neural differentiation was measured in three random microscopy fields (control [CTRL]: white bar; Rev-erbβ knockdown [Revβ-KD]: black bar). Error bars represent ± SEM (n = 4). P = 0.2582, P = 0.9856, P > 0.9999, P = 0.8715, Sidak’s multiple comparison test after two-way ANOVA
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Supplementary material 2 (PDF 1158 kb) Supplementary Material 2 Expression of representative neural stem cell (NSC) marker genes after Rev-erbβ gene knockdown in cultured adult NSCs. Total RNA from NSCs was analyzed by qRT-PCR after Rev-erbβ knockdown and puromycin selection (control [CTRL]: white bars; Rev-erbβ knockdown [Revβ-KD]: black bars). Data represent mean ± SD (n = 4). *P < 0.05, **P < 0.01, ****P ≤ 0.0001, two-tailed unpaired t test. N.D.: (not detected)
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Shimozaki, K. Involvement of Nuclear Receptor REV-ERBβ in Formation of Neurites and Proliferation of Cultured Adult Neural Stem Cells. Cell Mol Neurobiol 38, 1051–1065 (2018). https://doi.org/10.1007/s10571-018-0576-7
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DOI: https://doi.org/10.1007/s10571-018-0576-7