
==== Front
bioRxiv
BIORXIV
bioRxiv
2692-8205
Cold Spring Harbor Laboratory

10.1101/2024.08.28.610074
preprint
1
Article
O-glycosylation contributes to mammalian glycoRNA biogenesis
Porat Jennifer http://orcid.org/0000-0002-1709-4635

Watkins Christopher P. http://orcid.org/0009-0000-5045-5001

Jin Chunsheng
Xie Xixuan http://orcid.org/0000-0002-8512-6053

Tan Xiao
Lebedenko Charlotta G. http://orcid.org/0000-0002-9891-9247

Hemberger Helena http://orcid.org/0009-0007-4535-5859

Shin Woojung http://orcid.org/0000-0002-1780-0317

Chai Peiyuan http://orcid.org/0000-0002-9203-8975

Collins James J.
Garcia Benjamin A.
Bojar Daniel http://orcid.org/0000-0002-3008-7851

Flynn Ryan A. http://orcid.org/0000-0001-5013-0442

29 8 2024
2024.08.28.610074https://creativecommons.org/licenses/by-nc-nd/4.0/ This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License, which allows reusers to copy and distribute the material in any medium or format in unadapted form only, for noncommercial purposes only, and only so long as attribution is given to the creator.
http://biorxiv.org/lookup/doi/10.1101/2024.08.28.610074
nihpp-2024.08.28.610074.pdf
Abstract

There is an increasing appreciation for the role of cell surface glycans in modulating interactions with extracellular ligands and participating in intercellular communication. We recently reported the existence of sialoglycoRNAs, where mammalian small RNAs are covalently linked to N-glycans through the modified base acp 3 U and trafficked to the cell surface. However, little is currently known about the role for O-glycosylation, another major class of carbohydrate polymer modifications. Here, we use parallel genetic, enzymatic, and mass spectrometry approaches to demonstrate that O-linked glycan biosynthesis is responsible for the majority of sialoglycoRNA levels. By examining the O-glycans associated with RNA from cell lines and colon organoids we find known and previously unreported O-linked glycan structures. Further, we find that O-linked glycans released from small RNA from organoids derived from ulcerative colitis patients exhibit higher levels of sialylation than glycans from healthy organoids. Together, our work provides flexible tools to interrogate O-linked glycoRNAs (O-glycoRNA) and suggests that they may be modulated in human disease.
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