Deregulated expression and subcellular localization of CPSF6, a circRNA-binding protein, promote malignant development of esophageal squamous cell carcinoma

Shichao Guo; Guangchao Wang; Zitong Zhao; Dan Li; Yongmei Song; Qimin Zhan

doi:10.21147/j.issn.1000-9604.2022.01.02

Deregulated expression and subcellular localization of CPSF6, a circRNA-binding protein, promote malignant development of esophageal squamous cell carcinoma

Chin J Cancer Res. 2022 Feb 28;34(1):11-27. doi: 10.21147/j.issn.1000-9604.2022.01.02.

Authors

Shichao Guo¹, Guangchao Wang¹, Zitong Zhao¹, Dan Li¹, Yongmei Song¹, Qimin Zhan^{1

2}

Affiliations

¹ State Key Laboratory of Molecular Oncology, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100021, China.
² Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Laboratory of Molecular Oncology, Peking University Cancer Hospital & Institute, Beijing 100142, China.

PMID: 35355934
PMCID: PMC8913258
DOI: 10.21147/j.issn.1000-9604.2022.01.02

Abstract

Objective: Cleavage and polyadenylation specific factor 6 (CPSF6) has been documented as an oncoprotein in different types of cancer. However, functions of CPSF6 have not been investigated yet in esophageal squamous cell carcinoma (ESCC). Here, we aimed to investigate the potential clinical values and biological functions of CPSF6 in ESCC.

Methods: For determining the expression level of CPSF6 in ESCC patients, we analyzed published data, performed quantitative real-time polymerase chain reaction (RT-qPCR) and immunohistochemistry assays. Kaplan-Meier curves and log-rank tests were used for survival analyses. GO and KEGG analyses were done for CPSF6-related genes. Cell proliferation, colony formation and xenograft assays were conducted to verify the effects of CPSF6 on ESCC. In addition, cell cycle and apoptosis assays were also performed to manifest the functions of CPSF6 and circCPSF6. RNA pulldown and radioimmunoprecipitation (RIP) assays were used for confirming the interaction between circCPSF6 (hsa_circ_0000417) and CPSF6 protein. The regulatory relationship between CPSF6 protein and circCPSF6 was determined by RT-qPCR.

Results: We found that CPSF6 was upregulated in ESCC tissues and overexpression of cytoplasmic CPSF6 was associated with poor prognosis. GO and KEGG analyses suggested that CPSF6 could mainly affect cell division in ESCC. Further experiments manifested that CPSF6 promoted cell proliferation and colony formationin vitro. Xenograft assay showed that knockdown of CPSF6 significantly decreased tumor growth rate in vivo. Subsequently, we verified that depletion of CPSF6 led to cell cycle arrest and apoptosis. Finally, we validated that CPSF6, as a circRNA-binding protein, interacted with and regulated its circular isoform circCPSF6 (hsa_circ_0000417), of which depletion also resulted in cell cycle arrest and cell apoptosis in ESCC.

Conclusions: These findings gave us insight that overexpression of cytoplasmic CPSF6 protein is associated with poor prognosis in ESCC and CPSF6 may function as an oncoprotein, at least in part, through regulating circCPSF6 expression.

Keywords: CPSF6; cell proliferation; circCPSF6; circRNA-binding protein; esophageal squamous cell carcinoma; prognosis.