Enhanced O-GlcNAc modification induced by the RAS/MAPK/CDK1 pathway is required for SOX2 protein expression and generation of cancer stem cells

Masahiro Shimizu,Hiroshi Shibuya,Nobuyuki Tanaka

doi:10.1038/s41598-022-06916-y

Masahiro Shimizu, Hiroshi Shibuya + Show 1 more

Open Access

https://doi.org/10.1038/s41598-022-06916-y

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Abstract

Cancer stem cells (CSCs) have tumour initiation, self-renewal, and long-term tumour repopulation properties, and it is postulated that differentiated somatic cells can be reprogrammed to CSCs by oncogenic signals. We previously showed that oncogenic HRASV12 conferred tumour initiation capacity in tumour suppressor p53-deficient (p53−/−) primary mouse embryonic fibroblasts (MEFs) through transcription factor NF-κB-mediated enhancement of glucose uptake; however, the underlying mechanisms of RAS oncogene-induced CSC reprogramming have not been elucidated. Here, we found that the expression of the reprogramming factor SOX2 was induced by HRASV12 in p53−/− MEFs. Moreover, gene knockout studies revealed that SOX2 is an essential factor for the generation of CSCs by HRASV12 in mouse and human fibroblasts. We demonstrated that HRASV12-induced cyclin-dependent kinase 1 (CDK1) activity and subsequent enhancement of protein O-GlcNAcylation were required for SOX2 induction and CSC generation in these fibroblasts and cancer cell lines containing RAS mutations. Moreover, the CDK inhibitor dinaciclib and O-GlcNAcylation inhibitor OSMI1 reduced the number of CSCs derived from these cells. Taken together, our results reveal a signalling pathway and mechanism for CSC generation by oncogenic RAS and suggest the possibility that this signalling pathway is a therapeutic target for CSCs.

Highlights

Cancer stem cells (CSCs) have tumour initiation, self-renewal, and long-term tumour repopulation properties, and it is postulated that differentiated somatic cells can be reprogrammed to CSCs by oncogenic signals
These results suggest that the expression of SRYbox 2 (SOX2) as well as Krüppel-like factor 4 (KLF4) was induced by HRASV12 expression in p53−/−mouse embryonic fibroblasts (MEFs), and OCT4 was induced in the process of CSC reprogramming
The dedifferentiation of somatic cells into CSCs is thought to be induced by a reprogramming mechanism similar to that observed in the production of iPSCs59,60

Summary

Introduction

Cancer stem cells (CSCs) have tumour initiation, self-renewal, and long-term tumour repopulation properties, and it is postulated that differentiated somatic cells can be reprogrammed to CSCs by oncogenic signals. We previously showed that oncogenic HRASV12 conferred tumour initiation capacity in tumour suppressor p53-deficient (p53−/−) primary mouse embryonic fibroblasts (MEFs) through transcription factor NF-κB-mediated enhancement of glucose uptake; the underlying mechanisms of RAS oncogene-induced CSC reprogramming have not been elucidated. We demonstrated that HRASV12-induced cyclindependent kinase 1 (CDK1) activity and subsequent enhancement of protein O-GlcNAcylation were required for SOX2 induction and CSC generation in these fibroblasts and cancer cell lines containing RAS mutations. Direct O-GlcNAcylation of OCT4 and SOX2 has been shown to regulate pluripotency and reprogramming of embryonic stem c ells[20] In relation to these results, we found that IL-8 overexpression in colon and lung cancer cells enhanced O-GlcNAcylation, which was required for the production and maintenance of C SCs21. Oncogenic mutations in the RAS-RAF-MAPK pathway contribute to various malignant phenotypes that include invasion, metastasis, relapse, and angiogenesis[5]

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