Glycosylated RNAs (GlycoRNAs) are a newly discovered class of molecules involved in glycosylation. They have been found on the surfaces of various cell types, organisms, and species, further advancing the role of RNA in biological processes. Despite this, our knowledge of glycoRNAs and their functions remains limited, primarily due to the lack of effective methods to visualize and study these molecules in living organisms. Existing methods (such as RNA blotting, NGS, HPLC, and MS) have limitations in specificity, spatial resolution, or direct visualization on cell surfaces.
UT Austin researchers have designed a novel method called Sialic Acid Aptamer and RNA in situ Hybridization-mediated Proximity Ligation Assay (ARPLA). This patent-pending method will directly visualize glycoRNAs on the cell surface with high sensitivity and selectivity, while also providing both sequence and spatial information, unlike current methods.
ARPLA utilizes a dual-probe system: a sialic acid aptamer (single-stranded nucleic acids that act as affinity reagent) for glycan-binding and an RNA hybridization probe to target the RNA part of the glycoRNA. The key to ARPLA is the in-situ proximity ligation assay, which requires that these two probes be close to each other to generate a signal. This dual recognition-in-proximity prevents false positive signals.
ARPLA’s high sensitivity, selectivity, and its ability to image unlabeled glycoRNAs make it a powerful tool for investigating the roles of glycoRNAs in many biological processes and uncovering their relevance to diseases. It offers several competitive advantages:
ARPLA has shown promise in breast cancer research, immune cell differentiation, and response to lipopolysaccharide (LPS) stimulation.