Study of human congenital disorders of glycosylation using fission yeasts as a model organism

GALLO GIOVANNA LUCRECIA; VALKO, AYELEN; ETCHEGARAY, EMILIANA; D'ALESSIO CECILIA; ARAMBURU, SOFÍA IVANA; PARODI ARMANDO

Bariloche

Simposio; 34° International Specialized Symposium on Yeasts; 2018

Institución organizadora del 34° International Specialized Symposium on Yeasts

N-glycosylation is one of the most frequent post-translational modifications of proteins that enter the secretory pathway. It is catalyzed by the oligosaccharyltransferase (OST), which transfers the pre-built glycan Glc3Man9GlcNAc2 from a Dolichol-PP derivative to the sequon NXS/T of proteins entering the endoplasmic reticulum (ER). Glycans play a key role in protein folding and in the quality control of glycoprotein folding. Mutations in genes involved in the Dolichol-PP-glycan biosynthetic pathway or in the transfer reaction produce protein hypoglycosylation (sequons normally occupied with a glycan are empty) thus producing folding defects and Congenital Disorders of Glycosylation (CDG) Type I. Defects in the N-glycan processing that is produced after the transfer reaction do not produce protein hypoglycosylation but aberrant glycan structures present in CDG Type II. The fission yeast Schizosaccharomyces pombe shares with mammalian cells all the mechanisms of Dolichol-PP-glycan biosynthesis, glycan transfer and the initial steps of N-glycan processing. Moreover, a large amount of genetic and biochemical tools make this yeast an ideal organism to model the molecular basis of CDG type I and some CDG type II. We constructed 16 yeast mutants that synthesize all possible combinations of Dolichol-PP-glycan structures to be transferred to proteins in the ER. (Glc0-3Man0-9GlcNAc2), mimicking the defects produced in CDG Type I. Results of protein hypoglycosylation in the mutants, analyzed both by flow cytometry of a fluorescent biosensor and by western blot, showed the relative involvement of each Glc and Man residue in the transfer efficiency by OST. The usefulness of yeast as model organism to study basic cell glycobiology processes will be discussed.