Abstract
A recent ground-breaking study suggested that small RNA from mammalian cells can undergo N-glycan modifications (termed glycoRNA)(1). The discovery relied upon a metabolic glycan labeling strategy in combination with commonly used phase-separation-based RNA isolation. Following the reported procedure, here we likewise identify an N-glycosylated species in the RNA fraction. However, our results suggest that the reported RNase sensitivity of the glycosylated species depends on the specific RNA purification method. This suggests the possibility of copurifying unexpected RNase-insensitive N-glycoconjugates during glycoRNA isolation. The co-existence of two independent, yet highly similar molecular entities, complicates biochemical assays on glycoRNA and calls for more specific approaches for glycoRNA analysis. To address this, we propose a control experiment that can help distinguish genuine glycoRNA species from copurified glycoconjugates.