N-linked glycosylation enrichment for in-depth cell surface proteomics of diffuse large B-cell lymphoma subtypes.

Global analysis of lymphoma genome integrity and transcriptomes tremendously advanced our understanding of their biology. Technological advances in mass spectrometry-based proteomics promise to complete the picture by allowing the global quantification of proteins and their post-translational modifications. Here we use N-glyco FASP, a recently developed mass spectrometric approach using lectin-enrichment, in conjunction with a super-SILAC approach to quantify N-linked glycoproteins in lymphoma cells. From patient-derived diffuse large B-cell lymphoma cell lines, we mapped 2383 glycosites on 1321 protein groups, which were highly enriched for cell membrane proteins. This N-glyco subproteome alone allowed the segregation of the ABC from the GCB subtypes of diffuse large B-cell lymphoma, which before gene expression studies had been considered one disease entity. Encouragingly, many of the glycopeptides driving the segregation belong to proteins previously characterized as segregators in a deep proteome study of these subtypes (S. J. Deeb et al. MCP 2012 PMID 22442255). This conforms to the high correlation that we observed between the expression level of the glycosites and their corresponding proteins. Detailed examination of glycosites and glycoprotein expression levels uncovered, among other interesting findings, enrichment of transcription factor binding motifs, including known NF-kappa-B related ones. Thus, enrichment of a class of post-translationally modified peptides can classify cancer types as well as reveal cancer specific mechanistic changes.     «

Global analysis of lymphoma genome integrity and transcriptomes tremendously advanced our understanding of their biology. Technological advances in mass spectrometry-based proteomics promise to complete the picture by allowing the global quantification of proteins and their post-translational modifications. Here we use N-glyco FASP, a recently developed mass spectrometric approach using lectin-enrichment, in conjunction with a super-SILAC approach to quantify N-linked glycoproteins in lymphoma c...     »