Epigenetic Repressing of CPEB1 Enhances Malignant Progression by Reducing Chromatin Accessibility of CEBPB in Colorectal Cancer

Abstract

Background: Cytoplasmic polyadenylation element-binding protein 1 (CPEB1), a sequence-specific RNA-binding protein that regulates mRNA polyadenylation and translation, has been linked to cancer progression and metastasis. However, the methylation role of CPEB1 and its underlying mechanism in colorectal cancer remain unclear.

Methods:  We identified hypermethylation of the CPEB1 gene as a potential novel epigenetic biomarker for CRC by clinical samples and its function were determined by in vitro and in vivo cell biology experiments. Meanwhile, its underlying mechanism was identified by a dual-luciferase reporter, chromatin immunoprecipitation, DNA pull-down, electrophoretic mobility shift assay.

Findings: CPEB1 hypermethylation was characterized through bioinformatic analysis of data catalogued in global-genome DNA methylation database. Hypermethylation status of CPEB1 in CRC tumor tissues and para-cancerous control tissues from 104 Chinese Southern Han patients was further validated by performing targeted bisulfite sequencing analysis. Silencing of the CPEB1 gene as a consequence of DNA hypermethylation was confirmed in both CRC tissues and cell lines. Furthermore, we demonstrated that up-regulating expression of CPEB1 and its tumor suppressor functionality in CRC cells significantly ameliorated tumor growth, migration, invasion, tumorigenicity and development while incrementally restoring cell apoptosis both in vitro and in vivo.  Mechanismly, we mapped the transcription factor (TF) and its binding-site within the CPEB1 promoter region and validated CEBPB as TF of CPEB1 and its binding site at -993—-779 of CPEB1 promoter region and demonstrated that the hypermethylation status could reduce CEBPB transcript activator activity and chromatin accessibility.

Interpretation: Our results delineated how silencing of CPEB1 functionally via epigenetic alterations contributes to malignant transformation of CRC by demonstrating a role for DNA hypermethylation as the underlying mechanism mediating CEBPB-binding attenuation and gene silencing.

Funding Statement: The work was supported by the National Natural Science Foundation of China (grant No. 81974313), Jiangsu Province's Key Young Medicine Talents Program (grant No. QNRC2016688, QNRC2016469), Postdoctoral Science Foundation of China (grant No. 2019M651930) and Nantong People's Livelihood Science and Technology Plan (MS12018032).

Declaration of Interests: The authors declare that they have no competing interests.

Ethics Approval Statement: All animal experiments were approved by the Institutional Animal Care and Use Committee (IACUC) of the Affiliated Hospital of Nantong University.