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MPDU1 regulates CEACAM1 and cell adhesion in vitro and in vivo

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Abstract

N-linked glycosylation (NLG) is a co-translational modification that is essential for the folding, stability, and trafficking of transmembrane (TM) and secretory glycoproteins. Efficient NLG requires the stepwise synthesis and en bloc transfer of a 14-sugar carbohydrate known as a lipid-linked oligosaccharide (LLO). The genetics of LLO biosynthesis have been established in yeast and Chinese hamster systems, but human models of LLO biosynthesis are lacking. In this study we report that Kato III human gastric cancer cells represent a model of deficient LLO synthesis, possessing a homozygous deletion of the LLO biosynthesis factor, MPDU1. Kato III cells lacking MPDU1 have all the hallmarks of a glycosylation-deficient cell line, including altered sensitivity to lectins and the formation of truncated LLOs. Analysis of transcription using an expression microarray and protein levels using a proteome antibody array reveal changes in the expression of several membrane proteins, including the metalloprotease ADAM-15 and the cell adhesion molecule CEACAM1. Surprisingly, the restoration of MPDU1 expression in Kato III cells demonstrated a clear phenotype of increased cell-cell adhesion, a finding that was confirmed in vivo through analysis of tumor xenografts. These experiments also confirmed that protein levels of CEACAM-1, which functions in cell adhesion, is dependent on LLO biosynthesis in vivo. Kato III cells and the MPDU1-rescued Kato IIIM cells therefore provide a novel model to examine the consequences of defective LLO biosynthesis both in vitro and in vivo.

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Abbreviations

LLO :

Lipid-linked oligosaccharide

NLG :

N-linked glycosylation

ER :

Endoplasmic reticulum

CHO :

Chinese hamster ovary

CDG :

Congenital disorder of glycosylation

MPDU :

Mannose-P-dolichol utilization

Man :

Mannose

GlcNac :

N-acetylglucosamine

FACE :

Fluorophore-assisted carbohydrate electrophoresis

CNV :

Copy number variation

H&E :

Hematoxylin and eosin

PBS :

Phosphate-buffered sailne

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Funding

This work was supported by grants from the National Institutes of Health (5RO1CA172391) and the National Research Service Award (postdoctoral training grant #T32CA009259).

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Authors and Affiliations

  1. Department of Therapeutic Radiology, Yale School of Medicine, New Haven, CT, 06510, USA

    Daniel C. Bennett, Aurelie Cazet, Jon Charest & Joseph N. Contessa

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  1. Daniel C. Bennett
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  2. Aurelie Cazet
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  3. Jon Charest
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  4. Joseph N. Contessa
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Correspondence to Joseph N. Contessa.

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Bennett, D.C., Cazet, A., Charest, J. et al. MPDU1 regulates CEACAM1 and cell adhesion in vitro and in vivo. Glycoconj J 35, 265–274 (2018). https://doi.org/10.1007/s10719-018-9819-6

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  • DOI: https://doi.org/10.1007/s10719-018-9819-6

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