MiR-129-5p targets Wnt5a to block PKC/ERK/NF-\u03baB and JNK pathways in glioblastoma

Ailiang Zeng,Xin Jin,Rui Li,Jianxing Yin,Feng Shen,Zheng Wang,Wei Yan,Xu Zhou,Yongping You,Yan Li

doi:10.1038/s41419-018-0343-1

Abstract

Therapeutic application of microRNAs (miRNAs) in Wnt-driven glioma has been valuable; however, their specific roles and mechanisms have not been completely investigated. Real-time quantitative PCR (RT-qPCR) was used to analyse the expression of microRNA-129-5p (miR-129-5p) in human glioma samples. Cell-Counting Kit 8 (CCK-8), flow cytometry, EdU, angiogenesis, Transwell invasion, wound healing, in vitro 3D migration and neurosphere formation assays were employed to assess the role of miR-129-5p in glioblastoma multiforme (GBM) cells. Moreover, we performed the luciferase reporter assay and the RNA-ChIP (chromatin immunoprecipitation) assay to confirm whether Wnt5a was a direct target of miR-129-5p. We also confirmed the correlation between the expression profile of miR-129-5p and Wnt5a in glioma patients from the Chinese Glioma Genome Atlas (CGGA) and investigated the overall survival of GBM patients using two data sets, namely, TCGA and GSE16011, according to their Wnt5a expression status. MiR-129-5p expression levels were downregulated and inversely correlated with Wnt5a expression levels in CGGA glioma patients. Restored expression of miR-129-5p blocked GBM cell proliferation, invasion, migration, angiogenesis, neurosphere formation and resistance to temozolomide. We reported that miR-129-5p directly targeted Wnt5a in glioma. Furthermore, we observed that overexpression of miR-129-5p inhibited the expression of Wnt5a, thus blocking the protein kinase C(PKC)/ERK/NF-κB and JNK pathways. Inhibiting Wnt5a rescued the effects of miR-129-5p loss and increased Wnt5a expression was associated with reduced overall survival of GBM patients. We also demonstrated the inhibitory effect of miR-129-5p on tumour growth in GBM using an in vivo model. The miR-129-5p/Wnt5a-axis-mediated PKC/ERK/NF-κB and JNK pathways have therapeutic potential in GBM treatment.

Highlights

Gliomas represent the most common primary brain tumours in adults and glioblastoma multiforme (GBM) has been categorised as a WHO grade IV disease[1]
Our findings demonstrate that miR-129-5p acts as a tumour suppressor, mediating the inhibition of proliferation, invasion, migration, neurosphere formation, angiogenesis and TMZ resistance by suppressing its target, Wnt5a plasmid (Wnt5a)
Ror1/2 and Ryk act as receptors or coreceptors for Wnt5a, and Ror1/2 and Ryk expression positively correlates with Wnt5a expression in GBM 12,13,15

Summary

Introduction

Gliomas represent the most common primary brain tumours in adults and glioblastoma multiforme (GBM) has been categorised as a WHO grade IV disease[1]. MiR-129-5p has been demonstrated to be downregulated in multiple types of cancers[2,3,4,5,6,7,8,9]. MiR-129-5p was observed to be downregulated in GBM tissues compared to that in adjacent non-tumourous tissues[10]. Zeng et al Cell Death and Disease (2018)9:394. Explicit molecular mechanisms of miR-129-5p in GBM need to be fully explored in multiple GBM cell lineages, including primary GBM cells and GBM stem cells (GSCs)

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Results

Conclusion