Installation of O-glycan sulfation capacities in human HEK293 cells for display of sulfated mucins
(2022) In Journal of Biological Chemistry 298(2).- Abstract
The human genome contains at least 35 genes that encode Golgi sulfotransferases that function in the secretory pathway, where they are involved in decorating glycosaminoglycans, glycolipids, and glycoproteins with sulfate groups. Although a number of important interactions by proteins such as selectins, galectins, and sialic acid-binding immunoglobulin-like lectins are thought to mainly rely on sulfated O-glycans, our insight into the sulfotransferases that modify these glycoproteins, and in particular GalNAc-type O-glycoproteins, is limited. Moreover, sulfated mucins appear to accumulate in respiratory diseases, arthritis, and cancer. To explore further the genetic and biosynthetic regulation of sulfated O-glycans, here we expanded a... (More)
The human genome contains at least 35 genes that encode Golgi sulfotransferases that function in the secretory pathway, where they are involved in decorating glycosaminoglycans, glycolipids, and glycoproteins with sulfate groups. Although a number of important interactions by proteins such as selectins, galectins, and sialic acid-binding immunoglobulin-like lectins are thought to mainly rely on sulfated O-glycans, our insight into the sulfotransferases that modify these glycoproteins, and in particular GalNAc-type O-glycoproteins, is limited. Moreover, sulfated mucins appear to accumulate in respiratory diseases, arthritis, and cancer. To explore further the genetic and biosynthetic regulation of sulfated O-glycans, here we expanded a cell-based glycan array in the human embryonic kidney 293 (HEK293) cell line with sulfation capacities. We stably engineered O-glycan sulfation capacities in HEK293 cells by site-directed knockin of sulfotransferase genes in combination with knockout of genes to eliminate endogenous O-glycan branching (core2 synthase gene GCNT1) and/or sialylation capacities in order to provide simplified substrates (core1 Galβ1-3GalNAcα1-O-Ser/Thr) for the introduced sulfotransferases. Expression of the galactose 3-O-sulfotransferase 2 in HEK293 cells resulted in sulfation of core1 and core2 O-glycans, whereas expression of galactose 3-O-sulfotransferase 4 resulted in sulfation of core1 only. We used the engineered cell library to dissect the binding specificity of galectin-4 and confirmed binding to the 3-O-sulfo-core1 O-glycan. This is a first step toward expanding the emerging cell-based glycan arrays with the important sulfation modification for display and production of glycoconjugates with sulfated O-glycans.
(Less)
- author
- organization
- publishing date
- 2022-02-01
- type
- Contribution to journal
- publication status
- published
- subject
- in
- Journal of Biological Chemistry
- volume
- 298
- issue
- 2
- article number
- 101382
- publisher
- American Society for Biochemistry and Molecular Biology
- external identifiers
-
- pmid:34954141
- scopus:85123414538
- ISSN
- 0021-9258
- DOI
- 10.1016/j.jbc.2021.101382
- language
- English
- LU publication?
- yes
- id
- 142073c1-480a-46d4-b249-89c1416d0376
- date added to LUP
- 2022-04-06 15:56:21
- date last changed
- 2024-04-24 20:00:29
@article{142073c1-480a-46d4-b249-89c1416d0376,
abstract = {{<p>The human genome contains at least 35 genes that encode Golgi sulfotransferases that function in the secretory pathway, where they are involved in decorating glycosaminoglycans, glycolipids, and glycoproteins with sulfate groups. Although a number of important interactions by proteins such as selectins, galectins, and sialic acid-binding immunoglobulin-like lectins are thought to mainly rely on sulfated O-glycans, our insight into the sulfotransferases that modify these glycoproteins, and in particular GalNAc-type O-glycoproteins, is limited. Moreover, sulfated mucins appear to accumulate in respiratory diseases, arthritis, and cancer. To explore further the genetic and biosynthetic regulation of sulfated O-glycans, here we expanded a cell-based glycan array in the human embryonic kidney 293 (HEK293) cell line with sulfation capacities. We stably engineered O-glycan sulfation capacities in HEK293 cells by site-directed knockin of sulfotransferase genes in combination with knockout of genes to eliminate endogenous O-glycan branching (core2 synthase gene GCNT1) and/or sialylation capacities in order to provide simplified substrates (core1 Galβ1-3GalNAcα1-O-Ser/Thr) for the introduced sulfotransferases. Expression of the galactose 3-O-sulfotransferase 2 in HEK293 cells resulted in sulfation of core1 and core2 O-glycans, whereas expression of galactose 3-O-sulfotransferase 4 resulted in sulfation of core1 only. We used the engineered cell library to dissect the binding specificity of galectin-4 and confirmed binding to the 3-O-sulfo-core1 O-glycan. This is a first step toward expanding the emerging cell-based glycan arrays with the important sulfation modification for display and production of glycoconjugates with sulfated O-glycans.</p>}},
author = {{Sun, Lingbo and Konstantinidi, Andriana and Ye, Zilu and Nason, Rebecca and Zhang, Yuecheng and Büll, Christian and Kahl-Knutson, Barbro and Hansen, Lars and Leffler, Hakon and Vakhrushev, Sergey Y. and Yang, Zhang and Clausen, Henrik and Narimatsu, Yoshiki}},
issn = {{0021-9258}},
language = {{eng}},
month = {{02}},
number = {{2}},
publisher = {{American Society for Biochemistry and Molecular Biology}},
series = {{Journal of Biological Chemistry}},
title = {{Installation of O-glycan sulfation capacities in human HEK293 cells for display of sulfated mucins}},
url = {{http://dx.doi.org/10.1016/j.jbc.2021.101382}},
doi = {{10.1016/j.jbc.2021.101382}},
volume = {{298}},
year = {{2022}},
}