期刊
ANALYTICAL CHEMISTRY
卷 95, 期 30, 页码 11326-11334出版社
AMER CHEMICAL SOC
DOI: 10.1021/acs.analchem.3c01392
关键词
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Single-cell omics is crucial for understanding population heterogeneity, identifying unique features of individual cells, and discovering minority subpopulations of interest. Protein N-glycosylation is a major post-translational modification that plays important roles in various biological processes. However, comprehensive N-glycoproteome profiling for single cells has been challenging due to limited sample amount and incompatibility with available enrichment strategies.
Single-cell omics is critical in revealing populationheterogeneity,discovering unique features of individual cells, and identifying minoritysubpopulations of interest. As one of the major post-translationalmodifications, protein N-glycosylation plays crucialroles in various important biological processes. Elucidation of thevariation in N-glycosylation patterns at single-cellresolution may largely facilitate the understanding of their key rolesin the tumor microenvironment and immune therapy. However, comprehensive N-glycoproteome profiling for single cells has not beenachieved due to the extremely limited sample amount and incompatibilitywith the available enrichment strategies. Here, we have developedan isobaric labeling-based carrier strategy for highly sensitive intact N-glycopeptide profiling for single cells or a small numberof rare cells without enrichment. Isobaric labeling has unique multiplexingproperties, by which the total signal from all channelstriggers MS/MS fragmentation for N-glycopeptide identification,while the reporter ions provide quantitative information. In our strategy,a carrier channel using N-glycopeptides obtainedfrom bulk-cell samples significantly improved the totalsignal of N-glycopeptides and, therefore, promotedthe first quantitative analysis of averagely 260 N-glycopeptides from single HeLa cells. We further applied this strategyto study the regional heterogeneity of N-glycosylationof microglia in mouse brain and discovered region-specific N-glycoproteome patterns and cell subtypes. In conclusion,the glycocarrier strategy provides an attractive solution for sensitiveand quantitative N-glycopeptide profiling of single/rarecells that cannot be enriched by traditional workflows.
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