Site‐specific O‐glycosylation of human granulocyte/macrophage colony‐stimulating factor secreted by yeast and animal cells

Abstract

To compare the site specificity of O‐glycosylation in lower and higher eukaryotes, we expressed human granulocyte/macrophage colony‐stimulating factor (hGM‐CSF) in the yeast Saccharomyces cerevisiae and in COS‐1 cells. Analyses of specific hGM‐CSF mutants secreted by yeast led to the conclusion that efficient O‐glycosylation in yeast requires residues S9 and T10. However, only S9 is used as an attachment point for an extended O‐glycosyl chain in a 15.5‐kDa hGM‐CSF form. A 14.5‐kDa hGM‐CSF form, secreted by yeast, appears substituted by single mannosyl residues at both positions S9 and T10, indicating that O‐glycosylation at T10 inhibits extension of the O‐glycosyl chain attached to S9. As in yeast cells, the addition of O‐glycosyl chains to hGM‐CSF secreted by COS‐1 cells requires the presence of S9 and T10 residues. These results demonstrate that, inspite of different biosynthetic routes, the selection of O‐glycosylation sites is similar between lower and higher eukaryotes. Copyright © 1992, Wiley Blackwell. All rights reserved