Novel Approach to Enriching Glycosylated RNAs: Specific Capture of GlycoRNAs via Solid-Phase Chemistry

Ribonuclease (RNA) modifications can alter cellular functionandlead to differential immune responses by acting as discriminatorsbetween RNAs from different phyla. RNA glycosylation has recentlybeen observed at the cell surface, and its dysregulation in diseasemay change RNA functions. However, determining which RNA substratescan be glycosylated remains to be explored. Here, we develop a solid-phasechemoenzymatic method (SPCgRNA) for targeting glycosylated RNAs, bywhich glycosylated RNA substrates can be specifically recognized.We found the differential N-glycosylation of smallRNAs in hTERT-HPNE and MIA PaCa-2 cancer cells using SPCgRNA. RNA-Seqshowed that the changes in glyco-miRNAs prepared from SPCgRNA wereconsistent with those of traditional methods. The KEGG signaling pathwayanalysis revealed that differential miRNA glycosylation can affecttumor cell proliferation and survival. Further studies found thatNGI-1 significantly inhibited the proliferation, migration, and circulationof MIA PaCa-2 and promoted cell apoptosis. In addition, & beta;-1,4-galactosyltransferase1 (B4GALT1) not only affected the expression levelof glycosylated miRNAs hsa-miR-21-5p but also promoted cell apoptosisand inhibited the cell cycle possibly through the p53 signaling pathway, while B4GALT1 and p53 were also affected following the hsa-miR-21-5p increase. These resultssuggest that B4GALT1 may catalyze miRNAs glycosylation,which further promotes cancer cell progression.

Automated summary

RNA modifications, specifically glycosylation, can alter cellular function and lead to differential immune responses. A solid-phase chemoenzymatic method has been developed to specifically recognize glycosylated RNA substrates. The differential glycosylation of small RNAs in cancer cells has been observed, and it affects tumor cell proliferation and survival.