Absence of nucleoside diphosphatase activities in the yeast secretory pathway does not abolish nucleotide sugar-dependent protein glycosylation.
It is accepted that glycosyltransferase-generated nucleoside diphosphates are converted to monophosphates in the secretory pathway by nucleoside diphosphatases (NDPases) to provide substrates for antiport transport systems by which entrance of nucleotide sugars from the cytosol into the lumen is coupled to exit of nucleoside monophosphates. Working with Saccharomyces cerevisiae mutants affected in anterograde and/or retrograde endoplasmic reticulum (ER)-Golgi vesicular traffic and/or defective in one or both secretory pathway (Golgi) NDPases, we show that UDP-Glc: glycoprotein glucosyltransferase-mediated glucosylation is not dependent on the presence of NDPases or on ER-Golgi vesicular traffic and that GDP-Man-dependent N- and O-mannosylations are reduced but not abolished in the absence of NDPases in the secretory pathway. Further, the absence of the main Man-1-P transferase (a Golgi GMP-generating enzyme) does not modify the limited mannosylation observed in the absence of NDPases. Based on these results and on available additional information, we suggest that in the absence of NDPases, the already characterized nucleotide sugar transporters allow entrance of nucleotide sugars into the luminal compartments and that resulting nucleoside diphosphates exit to the cytosol by a still unknown mechanism. Further, an unexpected side result suggests that formation of Ser/Thr-Man(2) may occur in the ER and not exclusively in the Golgi.
D'Alessio
Cecilia
C
Laboratory of Glycobiology, Fundación Instituto Leloir, Avda. Patricias Argentinas 435, Buenos Aires C1405BWE, Argentina.
Caramelo
Julio J
JJ
Parodi
Armando J
AJ
D017766:Acid Anhydride Hydrolases / Q000502:physiology*; D001692:Biological Transport; D004721:Endoplasmic Reticulum / Q000378:metabolism; D015870:Gene Expression; D006031:Glycosylation / Q000187:drug effects*; D006056:Golgi Apparatus / Q000378:metabolism; D006595:Hexosamines / Q000378:metabolism; D006602:Hexosyltransferases / Q000235:genetics / Q000378:metabolism*; D006868:Hydrolysis; D009154:Mutation; D009702:Nucleoside Diphosphate Sugars / Q000378:metabolism; D009711:Nucleotides / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D012568:Schizosaccharomyces / Q000201:enzymology; D014162:Transfection; D014532:Uridine Diphosphate Glucose / Q000378:metabolism
D006595:Hexosamines; D009702:Nucleoside Diphosphate Sugars; D009711:Nucleotides; C008322:mannosamine; D006602:Hexosyltransferases; C020823:glycoprotein glycosyltransferase; D017766:Acid Anhydride Hydrolases; C023837:nucleoside-diphosphatase; D014532:Uridine Diphosphate Glucose