Total cellular glycomics allows characterizing cells and streamlining the discovery process for cellular biomarkers

TLDR

Pluripotency biomarkers are often glycoconjugates, yet discovering new glycoconjugate biomarkers is hindered by technical challenges. This study introduces a unique approach to systematically profile all major oligosaccharide classes in the cellular glycome. The method employs mass spectrometry to qualitatively and quantitatively analyze N‑ and O‑linked glycans, glycosaminoglycans, glycosphingolipids, and free oligosaccharides from hESCs, hiPSCs, and other human cells. The total glycomes proved highly cell‑specific, with hESCs and hiPSCs displaying immature, yet highly similar glycan profiles, and the technique identified known pluripotency markers and a panel of stem‑cell‑specific glycans.

Abstract

Although many of the frequently used pluripotency biomarkers are glycoconjugates, a glycoconjugate-based exploration of novel cellular biomarkers has proven difficult due to technical difficulties. This study reports a unique approach for the systematic overview of all major classes of oligosaccharides in the cellular glycome. The proposed method enabled mass spectrometry–based structurally intensive analyses, both qualitatively and quantitatively, of cellular N - and O -linked glycans derived from glycoproteins, glycosaminoglycans, and glycosphingolipids, as well as free oligosaccharides of human embryonic stem cells (hESCs), induced pluripotent stem cells (hiPSCs), and various human cells derived from normal and carcinoma cells. Cellular total glycomes were found to be highly cell specific, demonstrating their utility as unique cellular descriptors. Structures of glycans of all classes specifically observed in hESCs and hiPSCs tended to be immature in general, suggesting the presence of stem cell–specific glycosylation spectra. The current analysis revealed the high similarity of the total cellular glycome between hESCs and hiPSCs, although it was suggested that hESCs are more homogeneous than hiPSCs from a glycomic standpoint. Notably, this study enabled a priori identification of known pluripotency biomarkers such as SSEA-3, -4, and -5 and Tra-1–60/81, as well as a panel of glycans specifically expressed by hESCs and hiPSCs.