Persistence of three but not two Glc in glycoproteins produces severe defects in fission yeasts

ETCHEGARAY, E.; PARODI, A.J.; VALKO, A.; DALESSIO, C. ; ARAMBURU, S.I.; GALLO, G.L.

Buenos Aires

Simposio; 3rd Argentinian Symposium on Glycobiology; 2019

Glucosidase I (GI) is an endoplasmic reticulum (ER) membrane protein that removes the outermost Glc from Glc3Man9GlcNAc2 (G3M9) transferred to proteins. Deficiency of GI results in congenital disorder of glycosylation (CDG) IIb. In fission yeasts, lack of GI (Δgls1-S mutants) resulted in a very sick phenotype and accumulation of G3M9 protein-linked glycans. Surprisingly, mutants lacking glucosidase II, which display protein-linked G2M9, have a normal growth phenotype. This suggests that persistence of three but not two Glc in glycoproteins is deleterious for the cell. While searching Δgls1 mutants, we obtained a second strain with a healthier phenotype (Δgls1-H) that presented both G3M9 and Man9GlcNAc2 species. This strain had an additional spontaneous mutation in gene alg10 (adds outermost Glc during Dol-PP-G3M9 synthesis) thus producing N-linked G2M9, and bypassing the need for GI. Moreover, expression of Golgi human endomannosidase in Δgls1-S cells relieved the sick phenotype observed in this mutant, confirming that persistence of G3M9 species in glycoproteins is deleterious. As the phenotype reversion was not a complete one in either case we expressed two catalytic mutants of GI, resulting in a partial phenotype rescue. Altogether, these results would indicate that persistence of G3M9 in glycoproteins is at least in part responsible for the defects observed in the CDG-IIb disorder and also suggest that GI might have an additional function besides the catalytic one.