N6‐methyladenosine‐modified circTEAD1 stabilizes Yap1 mRNA to promote chordoma tumorigenesis

• Published in: Clinical and translational medicine 2024-04, Vol.14 (4), p.e1658-n/a
• Main Authors: Li, Hanwen, Tang, Yingchuang, Ruan, Xingbang, Zhang, Junxin, Liu, Hao, Yu, Shiyu, Chen, Hao, Yang, Huilin, Zhang, Kai, Chen, Kangwu
• Format: Article
• Language: English
• Subjects: Adaptor Proteins, Signal Transducing - genetics; Adaptor Proteins, Signal Transducing - metabolism; Adenosine - analogs & derivatives; Adenosine - genetics; Adenosine - metabolism; Animals; Bone cancer; Carcinogenesis - genetics; Cell Line, Tumor; Chordoma; Chordoma - genetics; Chordoma - metabolism; Chordoma - pathology; CircRNAs; Hippo pathway; Humans; Investigations; Localization; Medical prognosis; Mice; N6‐methyladenosine modification; Pathogenesis; Proteins; RNA, Circular - genetics; RNA, Circular - metabolism; RNA, Messenger - genetics; RNA, Messenger - metabolism; Transcription Factors - genetics; Transcription Factors - metabolism; Tumorigenesis; YAP-Signaling Proteins - genetics; YAP-Signaling Proteins - metabolism
• ISSN: 2001-1326; 2001-1326
• DOI: 10.1002/ctm2.1658

Background Chordoma, a rare bone tumour with aggressive local invasion and high recurrence rate with limited understanding of its molecular mechanisms. Circular RNAs (circRNAs) have been extensively implicated in tumorigenesis, yet their involvement in chordoma remains largely unexplored. N6‐methyladenosine (m6A) modification holds a crucial function in regulating protein translation, RNA degradation and transcription. Methods Initially, screening and validation of circTEAD1 in chordoma were conducted by high‐throughput sequencing. Subsequently, sh‐circTEAD1 and an overexpression plasmid were constructed. Colony formation assays, cell counting kit‐8, Transwell and wound healing assays were utilized to validate the function of circTEAD1 in vitro. RNA pull‐down assays identified the binding proteins of circTEAD1, which underwent verification through RNA immunoprecipitation (RIP). Methylated RIP assays were conducted to detect the m6A binding sites. Following this, luciferase assay, RT‐qPCR, RIP and Western blotting analyses were conducted, revealing that Yap1 was the direct target of circTEAD1. Afterwards, the same methods were utilized for the validation of the function of Yap1 in chordoma in vitro. Finally, the regulatory relationship between circTEAD1 and Yap1 in chordoma was verified by an in vivo tumour formation assay. Results CircTEAD1 was identified as an upregulated circRNA in chordoma specimens, with heightened circTEAD1 expression emerging as a prognostic indicator. In vitro experiments convincingly demonstrated that circTEAD1 significantly promoted chordoma cell invasion, migration and aggressiveness. Furthermore, the analysis revealed that methyltransferase‐like 3‐mediated m6A modification facilitated the cytoplasmic export of circTEAD1. The circTEAD1/IGF2BP3/Yap1 mRNA RNA‐protein ternary complex not only bolstered the stability of Yap1 mRNA but also exerted a pivotal role in driving chordoma tumorigenesis. Conclusions In this study, the role of m6A‐modified circTEAD1 in chordoma was identified. The findings offer novel insights into the potential molecular targets for chordoma therapy, shedding light on the intricate interplay between circRNAs, m6A modification and Yap1 mRNA in chordoma pathogenesis. Highlights: CircTEAD1 is an upregulated circRNA that serves as a prognostic indicator in chordoma. The export of circTEAD1 to the cytoplasm is mediated by METTL3 in an m6A methylation‐dependent manner. CircTEAD1 enhances Yap1 mRNA stability by form