Collaborative Research: Decoding GlycoRNAs on Cell Surfaces using Single-molecule and Single-cell Approaches

Researchers at the Rice University and University of Texas at Austin are developing new chemical biology tools to investigate glycosylated RNAs (glycoRNAs), a recently discovered class of small non-coding RNAs modified with glycans and displayed on the cell surface. GlycoRNAs have been identified across diverse cell types and species, indicating that they represent a conserved and biologically important class of biomolecules. Although glycosylated biomolecules regulate numerous essential biological processes, the fundamental biology and chemistry of glycoRNAs, such as their biosynthetic origins, plasma membrane organization, molecular interaction networks, and cell-state-dependent expression, remain largely unexplored. Progress in this field has been limited by the lack of technologies capable of studying glycoRNAs at single-molecule and single-cell resolution. To address these challenges, the research team will develop advanced imaging methods that simultaneously visualize glycoRNAs and their nonglycosylated RNA counterparts in living cells. These tools will enable quantitative comparisons of their abundance, spatial organization, membrane dynamics, and intracellular trafficking, and provide new insights into the biosynthetic pathways, membrane anchoring mechanisms, and selective glycosylation of small non-coding RNAs. The investigators will further employ genetic perturbation approaches to determine how glycoRNAs regulate cell-surface signaling pathways. Single-molecule tracking and super-resolution microscopy will be used to elucidate how glycoRNAs interact with RNA-binding proteins, assemble into nanoscale membrane clusters, and participate in signaling microdomains. In addition, the project will establish a single-cell glycoRNA-mRNA co-sequencing platform by selectively attaching sequencing adapters to glycoRNAs through a proximity-based labeling strategy. This approach will enable glycoRNAs to be profiled using standard 3′ single-cell RNA sequencing workflows, allowing glycoRNA expression to be directly integrated with transcriptomic data from thousands of individual cells. The resulting datasets will reveal cell-type-specific glycoRNA expression patterns and provide foundational insights into the biological functions and regulatory roles of glycoRNAs. Collectively, this research will address critical knowledge gaps in glycoRNA biology while creating broadly applicable technologies for studying glycoRNAs, RNA modifications, extracellular RNAs, and membrane-associated biomolecules. These advances are expected to establish glycoRNAs as a new class of molecular targets for disease diagnosis and therapeutic intervention and to enable future discoveries across chemical biology, glycobiology, RNA biology. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.