title: Mutation of N-linked glycosylation at Asn548 in CD133 decreases its ability to promote hepatoma cell growth.
abstract: The membrane glycoprotein CD133 is a popular marker for cancer stem cells and contributes to cancer initiation and invasion in a number of tumor types. CD133 promotes tumorigenesis partly through an interaction between its phosphorylated Y828 residue and the PI3K regulatory subunit p85, and the interaction with β-catenin. Although CD133 glycosylation is supposed to be associated with its function, the contribution of N-glycosylation to its functions remains unclear. Here we analyzed the exact site(s) of N-glycosylation in CD133 by mass spectrometry and found that all eight potential N-glycosylation sites of CD133 could be indeed occupied by N-glycans. Loss of individual N-glycosylation sites had no effect on the level of expression or membrane localization of CD133. However, mutation at glycosylation site Asn548 significantly decreased the ability of CD133 to promote hepatoma cell growth. Furthermore, mutation of Asn548 reduced the interaction between CD133 and β-catenin and inhibited the activation of β-catenin signaling by CD133 overexpression. Our results identified the characteristics and function of CD133 glycosylation sites. These data could potentially shed light on molecular regulation of CD133 by glycosylation and enhance our understanding of the utility of glycosylated CD133 as a target for cancer therapies.
authors.affiliation: Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Department of Biochemistry and Molecular Biology, Shanghai Medical College of Fudan University, Shanghai, People's Republic of China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Department of Biochemistry and Molecular Biology, Shanghai Medical College of Fudan University, Shanghai, People's Republic of China.
authors.forename: Shifang
authors.initials: S
authors.lastname: Ren
authors.affiliation: Institutes of Biomedical Sciences of Fudan University, Shanghai, People's Republic of China.
authors.forename: Liqi
authors.initials: L
authors.lastname: Xie
authors.affiliation: Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Department of Biochemistry and Molecular Biology, Shanghai Medical College of Fudan University, Shanghai, People's Republic of China.
authors.forename: Chunhong
authors.initials: C
authors.lastname: Cui
authors.affiliation: Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Department of Biochemistry and Molecular Biology, Shanghai Medical College of Fudan University, Shanghai, People's Republic of China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Xing
authors.affiliation: Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Department of Biochemistry and Molecular Biology, Shanghai Medical College of Fudan University, Shanghai, People's Republic of China.
authors.forename: Chanjuan
authors.initials: C
authors.lastname: Liu
authors.affiliation: Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Department of Biochemistry and Molecular Biology, Shanghai Medical College of Fudan University, Shanghai, People's Republic of China.
authors.forename: Benjin
authors.initials: B
authors.lastname: Cao
authors.affiliation: Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Department of Biochemistry and Molecular Biology, Shanghai Medical College of Fudan University, Shanghai, People's Republic of China.
authors.forename: Fan
authors.initials: F
authors.lastname: Yang
authors.affiliation: Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Department of Biochemistry and Molecular Biology, Shanghai Medical College of Fudan University, Shanghai, People's Republic of China.
authors.forename: Yinan
authors.initials: Y
authors.lastname: Li
authors.affiliation: Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Department of Biochemistry and Molecular Biology, Shanghai Medical College of Fudan University, Shanghai, People's Republic of China.
authors.forename: Xiaoning
authors.initials: X
authors.lastname: Chen
authors.affiliation: Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Department of Biochemistry and Molecular Biology, Shanghai Medical College of Fudan University, Shanghai, People's Republic of China.
authors.forename: Yuanyan
authors.initials: Y
authors.lastname: Wei
authors.affiliation: Institutes of Biomedical Sciences of Fudan University, Shanghai, People's Republic of China.
authors.forename: Haojie
authors.initials: H
authors.lastname: Lu
authors.affiliation: Key Laboratory of Glycoconjuates Research, Ministry of Public Health and Gene Research Center, Department of Biochemistry and Molecular Biology, Shanghai Medical College of Fudan University, Shanghai, People's Republic of China.
authors.forename: Jianhai
authors.initials: J
authors.lastname: Jiang
mesh_terms: D000071916:AC133 Antigen; D000595:Amino Acid Sequence; D015703:Antigens, CD / Q000235:genetics* / Q000378:metabolism*; D001216:Asparagine / Q000235:genetics / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000235:genetics* / Q000473:pathology; D048708:Cell Growth Processes / Q000235:genetics; D006023:Glycoproteins / Q000235:genetics* / Q000378:metabolism*; D006031:Glycosylation; D056945:Hep G2 Cells; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics* / Q000473:pathology; D008969:Molecular Sequence Data; D009154:Mutation; D010455:Peptides / Q000235:genetics* / Q000378:metabolism*
chemical_list: D000071916:AC133 Antigen; D015703:Antigens, CD; D006023:Glycoproteins; C000606023:PROM1 protein, human; D010455:Peptides; D001216:Asparagine
keywords: CD133; N-linked glycosylation; cell growth; hepatoma