SMURF2 phosphorylation at Thr249 modifies glioma stemness and tumorigenicity by regulating TGF-\u03b2 receptor stability

Manami Hiraiwa,Kazuya Tokumura,Mitsutoshi Nakada,Atsushi Hirao,Gyujin Park,Takashi Iezaki,Katsuyuki Kaneda,Sayuki Iwahashi,Tomoki Todo,Kazuya Fukasawa,Masaki Kobayashi,Hemragul Sabit,Tetsuhiro Horie,Misato Murata,Akane Suzuki,Eiichi Hinoi

doi:10.1038/s42003-021-02950-0

Abstract

Glioma stem cells (GSCs) contribute to the pathogenesis of glioblastoma, the most malignant form of glioma. The implication and underlying mechanisms of SMAD specific E3 ubiquitin protein ligase 2 (SMURF2) on the GSC phenotypes remain unknown. We previously demonstrated that SMURF2 phosphorylation at Thr249 (SMURF2Thr249) activates its E3 ubiquitin ligase activity. Here, we demonstrate that SMURF2Thr249 phosphorylation plays an essential role in maintaining GSC stemness and tumorigenicity. SMURF2 silencing augmented the self-renewal potential and tumorigenicity of patient-derived GSCs. The SMURF2Thr249 phosphorylation level was low in human glioblastoma pathology specimens. Introduction of the SMURF2T249A mutant resulted in increased stemness and tumorigenicity of GSCs, recapitulating the SMURF2 silencing. Moreover, the inactivation of SMURF2Thr249 phosphorylation increases TGF-β receptor (TGFBR) protein stability. Indeed, TGFBR1 knockdown markedly counteracted the GSC phenotypes by SMURF2T249A mutant. These findings highlight the importance of SMURF2Thr249 phosphorylation in maintaining GSC phenotypes, thereby demonstrating a potential target for GSC-directed therapy.

Highlights

Glioma stem cells (GSCs) contribute to the pathogenesis of glioblastoma, the most malignant form of glioma
We recently demonstrated that the phosphorylation of SMURF2 at Thr[249] (SMURF2Thr249) by extracellular signal-regulated kinase 5 (ERK5) plays an essential role in maintaining the stemness of mesenchymal stem cells (MSCs), which contributes to skeletogenesis[10]
Ubiquitination of endogenous TGFBR1 and TGFBR2 was markedly elevated by the overexpression of SMURF2WT, but it was decreased by the enforced infection of SMURF2T249A in TGS-01 GSCs (Fig. 4d). These results suggest that SMURF2Thr[249] phosphorylation decreases the protein stability of TGFBR by enhancing its E3 ubiquitin ligase activity, which in turn reduced Transforming growth factor-β (TGF-β)-SMAD2/3 signaling to repress the self-renewal potential and tumorigenicity of GSCs

Summary

Introduction

Glioma stem cells (GSCs) contribute to the pathogenesis of glioblastoma, the most malignant form of glioma. TGFBR1 knockdown markedly counteracted the GSC phenotypes by SMURF2T249A mutant These findings highlight the importance of SMURF2Thr[249] phosphorylation in maintaining GSC phenotypes, thereby demonstrating a potential target for GSC-directed therapy. We recently demonstrated that the phosphorylation of SMURF2 at Thr[249] (SMURF2Thr249) by extracellular signal-regulated kinase 5 (ERK5) plays an essential role in maintaining the stemness of mesenchymal stem cells (MSCs), which contributes to skeletogenesis[10]. SMURF2Thr[249] phosphorylation activates its E3 ubiquitin ligase activity, which modifies the stability of SMAD proteins, which in turn transcriptionally activate the expression of SOX9, the principal transcription factor of skeletogenesis in MSCs. Gliomas, which represent ~80% of all primary malignant brain tumors in humans, can be categorized into four grades according to the World Health Organization (WHO) classification criteria: grade I, grade II, grade III, and grade IV (glioblastoma, GBM)[11,12]. Recent studies have determined that GSCs contribute to high rates of therapeutic resistance and rapid recurrence[18,19], cancer invasion, immune evasion, tumor angiogenesis, and the recruitment of tumor-associated macrophages, which indicates that targeting GSCs is an efficacious strategy for improving GBM treatment[20,21,22]

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Conclusion