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

VALKO, AYELÉN; D'ALESSIO, C; ARAMBURU, SOFÍA I.; GALLO, GIOVANNA L; ETCHEGARAY, EMILIANA; PARODI AJ

Partido de San Martín, Buenos Aires

Congreso; 3rd Argentinean Symposium on G; 2019

Universidad Nacional de San Martín

Glucosidase I(GI) is an endoplasmic reticulum (ER) membrane protein that removes theoutermost Glc from Glc3Man9GlcNAc2 (G3M9)transferred to proteins. Deficiency of GI results in congenital disorder ofglycosylation (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 lackingglucosidase II, whichdisplay protein-linked G2M9, have a normal growth phenotype. This suggeststhat persistence of three but not two Glc in glycoproteins is deleterious forthe cell. While searching Δgls1 mutants, we obtained a second strainwith a healthier phenotype (Δgls1-H) that presented both G3M9 and Man9GlcNAc2species. Thisstrain had an additional spontaneous mutation in gene alg10 (addsoutermost Glc during Dol-PP-G3M9 synthesis) thus producing N-linked G2M9,and bypassing the need for GI. Moreover, expression of Golgi humanendomannosidase in Δgls1-S cells relieved the sick phenotype observed inthis mutant, confirming that persistence of G3M9 species in glycoproteins isdeleterious. As the phenotype reversion was not a complete one in either casewe expressed two catalytic mutants of GI, resulting in a partial phenotyperescue. Altogether, these results would indicate that persistence of G3M9 inglycoproteins is at least in part responsible for the defects observed in theCDG-IIb disorder and also suggest that GI might have an additional functionbesides the catalytic one.