O-Glycosylated RNA Identification and Site-specific Prediction by Solid-phase Chemoenzymatic TnORNA method and PONglyRNA tool

作者：Jiajia Li, Linshu Wang, Yan Chen, Shaomei Zhang, Zhongmin Wen, Xuechu Zhen, Haiyun Zhang, Yuan Zhou, Longjiang Xu, Shuang Yang · 发表于：bioRxiv (Cold Spring Harbor Laboratory) · 年份：2024 · DOI：10.1101/2024.06.18.599663 · 被引用次数：5 · 研究领域：RNA and protein synthesis mechanisms、Molecular Biology Techniques and Applications、RNA modifications and cancer

Abstract Recent studies have shown that the cell surface undergoes post-transcriptional modification by N-linked glycosylation. However, the question of whether RNA can be glycosylated by O-glycans remains to be explored. The presence of O-glycosylation in cells is indirectly revealed by the presence of O-glycans on RNAs following treatment with O-glycoproteases. To identify RNA O-glycosylation, we have developed a chemoenzymatic method for capturing and enriching O-glycosylated RNA (O-glycoRNA) using covalent immobilization on a solid support. GalNAcEXO selectively releases Tn-containing O-glycosylated RNAs (TnORNA). Using this method and SPCgRNA, we compared the expression of O-glycoRNAs and N-glycoRNAs in pancreatic cancer cell lines and tissues. We found that glycosylated miR-103a-3p, miR-122-5p, and miR-4492 regulate pancreatic cancer cell growth and proliferation through the PI3K-Akt pathway. In vitro assays and PDAC tissue analysis confirmed the potential regulatory roles of Tn-O-glycosylated miRNAs in pancreatic tumor growth and metastasis. Furthermore, a significant number (131) of miRNAs carrying both N- and Tn-O-glycosylation were identified, indicating the co-occurrence of N-linked and O-linked glycosylation on small RNAs. We have also developed PONglyRNA, an online bioinformatic tool for the site-specific prediction of RNA glycosylation. PONglyRNA identifies glycosylation motifs based on RNA sequence and has been validated using our glycoRNA data. In conclusion, ...