A New Type of Congenital Disorders of Glycosylation (CDG-Ii) Provides New Insights into the Early Steps of Dolichol-linked Oligosaccharide Biosynthesis

Christian Thiel,Markus Schwarz,Jianhe Peng,Michal Grzmil,Martin Hasilik,Thomas Braulke,Alfried Kohlschütter,Kurt Von Figura,Ludwig Lehle,Christian Körner

doi:10.1074/jbc.m302850200

Abstract

Deficiency of GDP-Man:Man1GlcNAc2-PP-dolichol mannosyltransferase (hALG2), is the cause of a new type of congenital disorders of glycosylation (CDG) designated CDG-Ii. The patient presented normal at birth but developed in the 1st year of life a multisystemic disorder with mental retardation, seizures, coloboma of the iris, hypomyelination, hepatomegaly, and coagulation abnormalities. An accumulation of Man1GlcNAc2-PP-dolichol and Man2GlcNAc2-PP-dolichol was observed in skin fibroblasts of the patient. Incubation of patient fibroblast extracts with Man1GlcNAc2-PP-dolichol and GDP-mannose revealed a severely reduced activity of the mannosyltransferase elongating Man1GlcNAc2-PP dolichol. Because the Saccharomyces cerevisiae mutant alg2-1 was known to accumulate the same shortened dolichol-linked oligosaccharides as the patient, the yeast ALG2 sequence was used to identify the human ortholog. Genetic analysis revealed that the patient was heterozygous for a single nucleotide deletion and a single nucleotide substitution in the human ortholog of yeast ALG2. Expression of wild type but not of mutant hALG2 cDNA restored the mannosyltransferase activity and the biosynthesis of dolichol-linked oligosaccharides both in patient fibroblasts and in the alg2-1 yeast cells. hALG2 was shown to act as an alpha1,3-mannosyltransferase. The resulting Manalpha1,3-ManGlcNAc2-PP dolichol is further elongated by a yet unknown alpha1,6-mannosyltransferase.

Highlights

Deficiency of GDP-Man:Man1GlcNAc2-PP-dolichol mannosyltransferase, is the cause of a new type of congenital disorders of glycosylation (CDG) designated CDG-Ii
Congenital disorders of glycosylation (CDG)1 compose a rapidly growing group of inherited multisystemic disorders in man, which are commonly associated with severe psychomotor and mental retardation [1]
We show that the affected mannosyltransferase is the human ortholog to the yeast ALG2 gene, an enzyme that has so far not been characterized in higher eukaryotes

Summary

Introduction

Deficiency of GDP-Man:Man1GlcNAc2-PP-dolichol mannosyltransferase (hALG2), is the cause of a new type of congenital disorders of glycosylation (CDG) designated CDG-Ii. Because the Saccharomyces cerevisiae mutant alg was known to accumulate the same shortened dolichol-linked oligosaccharides as the patient, the yeast ALG2 sequence was used to identify the human ortholog. Expression of wild type but not of mutant hALG2 cDNA restored the mannosyltransferase activity and the biosynthesis of dolichol-linked oligosaccharides both in patient fibroblasts and in the alg yeast cells. We describe for the first time a molecular defect in glycoprotein biosynthesis in man which affects at the cytosolic side of the endoplasmic reticulum the transfer of mannosyl residues from GDP-Man to Man1GlcNAc2-PP-dolichol by the enzyme hALG2. The characterization of the human ALG2 deficiency described here has helped to define ALG2, both from man and yeast, as the ␣1,3-mannosyltransferase that catalyzes the transfer of mannose residues onto Man1GlcNAc2-PP-dolichol

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Conclusion