title: The analysis of N-glycans of cell membrane proteins from human hematopoietic cell lines reveals distinctions in their pattern.
abstract: Human cell lines are often different in their features and present variations in the glycosylation patterns of cell membrane proteins. Protein glycosylation is the most common posttranslational modification and plays a particular role in functionality and bioactivity. The key approach of this study is the comparative analysis of five hematopoietic cell lines for their N-glycosylation pattern. The N-glycans of membrane proteins were elucidated by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and MALDI-TOF/TOF-MS analyses. Furthermore, the expression of a set of glycosyltransferases was determined via RT-PCR. The B-lymphoma BJA-B and promyelocytic HL-60 cell lines distinguish in levels and linkages of glycan-bound sialic acids. Furthermore, subclones of BJA-B and HL-60 cells, which completely lack UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE), the key enzyme of sialic acid biosynthesis, contained almost no sialylated N-glycans. Compared to wild-type cells, the GNE-deficient cells presented a similar cell surface N-glycosylation pattern in terms of antennarity and fucosylation. The Jurkat T-cell line revealed only partially sialylated N-glycans. Additionally, the different hematopoietic cell lines vary in their level of bisecting GlcNAcylation and antennary fucosylation with the quantities of bisecting N-acetylglucosamine (GlcNAc) and core fucose coinciding with the expression of GnT-III and FucT-VIII. Of note is the occurrence of N-acetyllactosamine (LacNAc) extensions on tetraantennary structures in GNE-deficient cell lines.
authors.affiliation: Laboratory of Biochemistry, Department of Life Sciences and Technology, University of Applied Sciences Berlin, Seestrasse 64, 13347 Berlin, Germany. stefan.reinke@beuth-hochschule.de
authors.forename: Stefan O
authors.initials: SO
authors.lastname: Reinke
authors.forename: Marion
authors.initials: M
authors.lastname: Bayer
authors.forename: Markus
authors.initials: M
authors.lastname: Berger
authors.forename: Stephan
authors.initials: S
authors.lastname: Hinderlich
authors.forename: Véronique
authors.initials: V
authors.lastname: Blanchard
mesh_terms: D000606:Amino Sugars / Q000032:analysis; D001773:Blood Cells / Q000737:chemistry*; D002240:Carbohydrate Sequence; D002460:Cell Line; D006031:Glycosylation; D016695:Glycosyltransferases / Q000235:genetics; D018922:HL-60 Cells; D006412:Hematopoietic Stem Cells / Q000737:chemistry*; D006801:Humans; D019169:Jurkat Cells; D008565:Membrane Proteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics; D011134:Polysaccharides / Q000032:analysis*; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D000606:Amino Sugars; D008565:Membrane Proteins; D011134:Polysaccharides; D012333:RNA, Messenger; C000458:N-acetyllactosamine; D016695:Glycosyltransferases; D017853:Phosphotransferases (Alcohol Group Acceptor); C018765:N-acylmannosamine kinase