Abstract
Disruption of contact inhibition and serum afflux that occur after a tissue injury activate cell cycle, which then stops when confluence is reached again. Although the events involved in cell cycle entry have been widely documented, those managing cell cycle exit have remained so far ill defined. We have identified that the final stage of wound closure is preceded in keratinocytes by a strong accumulation of miR-483-3p, which acts as a mandatory signal triggering cell cycle arrest when confluence is reached. Blocking miR-483-3p accumulation strongly delays cell cycle exit, maintains cells into a proliferative state and retards their differentiation program. Using two models of cell cycle synchronization (i.e. mechanical injury and serum addition), we show that an ectopic upregulation of miR-483-3p blocks cell cycle progression in early G1 phase. This arrest results from a direct targeting of the CDC25A phosphatase by miR-483-3p, which can be impeded using an anti-miRNA against miR-483-3p or a protector that blocks the complex formation between miR-483-3p and the 3′-untranslated region (UTR) of CDC25A transcript. We show that the miRNA-induced silencing of CDC25A increases the tyrosine phosphorylation status of CDK4/6 cyclin-dependent kinases which, in turn, abolishes CDK4/6 capacity to associate with D-type cyclins. This prevents CDK4/6 kinases’ activation, impairs downstream events such as cyclin E stimulation and sequesters cells in early G1. We propose this new regulatory process of cyclin-CDK association as a general mechanism coupling miRNA-mediated CDC25A invalidation to CDK post-transcriptional modifications and cell cycle control.
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Abbreviations
- CDK:
-
cyclin-dependent kinase
- CKI:
-
CDK inhibitor
- FCS:
-
fetal calf serum
- KRT:
-
keratin
- LNA:
-
locked nucleic acid
- LOI:
-
loss of imprinting
- miRNA:
-
microRNA
- mRNA:
-
messenger RNA
- NHK:
-
Normal human keratinocytes
- NICD:
-
Notch intracellular domain
- siRNA:
-
short interfering RNA
- UTR:
-
untranslated region
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Acknowledgements
This work was supported by funds from the Association pour la Recherche sur le Cancer (ARC, Grant 5420). Thomas Bertero was a recipient of a doctoral contract from the Ministry of Higher Education and Research. Cécile Gastaldi was a recipient of a fellowship from INSERM/PACA region.
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Bertero, T., Gastaldi, C., Bourget-Ponzio, I. et al. CDC25A targeting by miR-483-3p decreases CCND–CDK4/6 assembly and contributes to cell cycle arrest. Cell Death Differ 20, 800–811 (2013). https://doi.org/10.1038/cdd.2013.5
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DOI: https://doi.org/10.1038/cdd.2013.5
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