Celastrol suppresses human pancreatic cancer via m6A-YTHDF3-mediated downregulation of Claspin and Bcl-2

Yang Zhou; Haoran Zhuang; Yuxiang Liu; Jing Yin; Xiaoying Wei; Yue Qiu; Zhen Tian; Tingyu Miao; Jing Chen; Peifen Li; Xiao Xu; Wenjuan Wu; Huanan Li; Weigan Shen

doi:10.1007/s12672-023-00838-5

Celastrol suppresses human pancreatic cancer via m⁶A-YTHDF3-mediated downregulation of Claspin and Bcl-2

Discov Oncol. 2023 Dec 18;14(1):233. doi: 10.1007/s12672-023-00838-5.

Authors

Yang Zhou^#¹, Haoran Zhuang^#¹, Yuxiang Liu^#¹, Jing Yin¹, Xiaoying Wei¹, Yue Qiu¹, Zhen Tian¹, Tingyu Miao¹, Jing Chen¹, Peifen Li¹, Xiao Xu², Wenjuan Wu³, Huanan Li⁴, Weigan Shen^{5

6}

Affiliations

¹ Department of Cell Biology, School of Medicine of Yangzhou University, Yangzhou, Jiangsu, China.
² Department of Oncology, Taizhou Hospital of Traditional Chinese Medicine, Taizhou, Jiangsu, China.
³ Department of Oncology, Clinical Medical College, Yangzhou University, Yangzhou, Jiangsu, China. wuwenjuanyzu@126.com.
⁴ Department of Cell Biology, School of Medicine of Yangzhou University, Yangzhou, Jiangsu, China. hnli@yzu.edu.cn.
⁵ Department of Cell Biology, School of Medicine of Yangzhou University, Yangzhou, Jiangsu, China. shenweigan@hotmail.com.
⁶ Jiangsu Key Laboratory of Integrated Traditional Chinese and Western Medicine for Prevention and Treatment of Senile Diseases, Yangzhou University, Yangzhou, Jiangsu, China. shenweigan@hotmail.com.

^# Contributed equally.

Abstract

Background: Celastrol has been revealed to exhibit anticancer pharmacological activity, however, the molecular mechanisms of celastrol involved in pancreatic cancer remain to be further elucidated. The present study was to illustrate whether celastrol suppresses pancreatic cancer through modulating RNA m⁶A modification.

Methods: Effect of celastrol treatment on the malignant phenotypes of pancreatic cancer cells was evaluated by CCK-8 assay, EdU assay, colony formation assay, flow cytometry analysis and subcutaneous xenograft experiments. RNA sequencing (RNA-seq) analysis was carried out to analyze the genes differentially expressed in celastrol-treated pancreatic cancer cells. RT-qPCR, Western blotting and immunohistochemistry were employed to evaluate the expression of the indicated genes. RNA dot blot and quantification of total RNA m⁶A modification assays, MeRIP-qPCR assay, RIP-qPCR assay, RNA stability and protein stability assays were applied to evaluate the regulatory mechanism of celastrol treatment in pancreatic cancer cells.

Results: We demonstrated that celastrol suppressed cell proliferation and induced cell cycle arrest and apoptosis of pancreatic cancer cells in vitro, and decreased tumor growth in vivo. Specifically, Bcl-2, Claspin, METTL3 and YTHDF3 were identified as the potential targets of celastrol treatment in pancreatic cancer cells. Moreover, our results indicated that celastrol treatment downregulated METTL3 and decreased m⁶A levels of Claspin and Bcl-2 mRNA, leading to the degradation of Claspin and Bcl-2 mRNA in pancreatic cancer cells. Furthermore, we revealed that celastrol treatment downregulated Claspin and Bcl-2, at least in part, in an m⁶A-YTHDF3-mediated manner in pancreatic cancer cells.

Conclusion: Our study highlighted a novel mechanism underlying celastrol-induced cellular proliferation inhibition and apoptosis in pancreatic cancer cells via m⁶A-YTHDF3-mediated downregulation of Claspin and Bcl-2.

Keywords: Celastrol; Claspin; Pancreatic cancer; RNA m6A modification.

Abstract

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