Abstract
Wnt/β-catenin signaling pathway plays essential roles in mammalian development and tissue homeostasis. MicroRNAs (miRNAs) are a class of regulators involved in modulating this pathway. In this study, we screened miRNAs regulating Wnt/β-catenin signaling by using a TopFlash based luciferase reporter. Surprisingly, we found that miR-142 inhibited Wnt/β-catenin signaling, which was inconsistent with a recent study showing that miR-142-3p targeted Adenomatous Polyposis Coli (APC) to upregulate Wnt/β-catenin signaling. Due to the discordance, we elaborated experiments by using extensive mutagenesis, which demonstrated that the stem-loop structure was important for miR-142 to efficiently suppress Wnt/β-catenin signaling. Moreover, the inhibitory effect of miR-142 relies on miR-142-3p rather than miR-142-5p. Further, we found that miR-142-3p directly modulated translation of Ctnnb1 mRNA (encoding β-catenin) through binding to its 3’ untranslated region (3’ UTR). Finally, miR-142 was able to repress cell cycle progression by inhibiting active Wnt/β-catenin signaling. Thus, our findings highlight the inhibitory role of miR-142-3p in Wnt/β-catenin signaling, which help to understand the complex regulation of Wnt/β-catenin signaling.
Highlights
Wnt/β-catenin signaling is an ancient, highly conserved pathway that plays pivotal roles in a wide variety of developmental and self-renewing processes
Wnt/βcatenin signaling is involved in the self-renewal of hematopoietic stem cells (HSCs), development and differentiation of T cells, B cells and dendritic cells (DC) [1], and whose abnormity is highly associated with tumorigenesis, such as leukemia. β-catenin, the key transcriptional coactivator of the Wnt/β-catenin pathway, is trapped and degraded in the cytoplasm by a destruction complex
The luciferase activities of pGL3-TopFlash reporter were increased upon Wnt3a stimulation in a dose-dependent manner, whereas such effects were blocked by miR-142-expressing-stable NIH3T3 cell lines (miR-142) (Fig 1E). These results indicated that miR-142 was a potential suppressor of Wnt/β-catenin signaling, which is inconsistent with a recently study reported that miR-142-3p activated the canonical WNT signaling pathway in human breast cancer stem cells through suppression of Adenomatous Polyposis Coli (APC) [16]
Summary
Wnt/β-catenin signaling is an ancient, highly conserved pathway that plays pivotal roles in a wide variety of developmental and self-renewing processes. Wnt/βcatenin signaling is involved in the self-renewal of hematopoietic stem cells (HSCs), development and differentiation of T cells, B cells and dendritic cells (DC) [1], and whose abnormity is highly associated with tumorigenesis, such as leukemia. Β-catenin, the key transcriptional coactivator of the Wnt/β-catenin pathway, is trapped and degraded in the cytoplasm by a destruction complex (encompassing GSK3β, Axin and APC). Upon activation by Wnt ligands, the subsequent phosphorylation events lead to inhibition of the APC complex-mediated degradation of β-catenin, facilitating β-catenin to accumulate and localize to the nucleus to form complex with the T-cell factor/lymphoid enhancer factor (TCF/LEF), and initiating the expression of Wnt target genes [2].
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