Timo Imbach

Deficiency of dolichol-phosphate-mannose synthase-1 causes congenital disorder of glycosylation type Ie

Congenital disorders of glycosylation (CDG), formerly known as carbohydrate-deficient glycoprotein syndromes, lead to diseases with variable clinical pictures. We report the delineation of a novel type of CDG identified in 2 children presenting with severe developmental delay, seizures, and dysmorphic features. We detected hypoglycosylation on serum transferrin and cerebrospinal fluid beta-trace protein. Lipid-linked oligosaccharides in the endoplasmic reticulum of patient fibroblasts showed an accumulation of the dolichyl pyrophosphate Man(5)GlcNAc(2) structure, compatible with the reduced dolichol-phosphate-mannose synthase (DolP-Man synthase) activity detected in these patients. Accordingly, 2 mutant alleles of the DolP-Man synthase DPM1 gene, 1 with a 274C>G transversion, the other with a 628delC deletion, were detected in both siblings. Complementation analysis using DPM1-null murine Thy1-deficient cells confirmed the detrimental effect of both mutations on the enzymatic activity. Furthermore, mannose supplementation failed to improve the glycosylation status of DPM1-deficient fibroblast cells, thus precluding a possible therapeutic application of mannose in the patients. Because DPM1 deficiency, like other subtypes of CDG-I, impairs the assembly of N-glycans, this novel glycosylation defect was named CDG-Ie.

Clinical and biochemical characteristics of congenital disorder of glycosylation type Ic, the first recognized endoplasmic reticulum defect in N-glycan synthesis

We report on 8 patients with a recently described novel subtype of congenital disorder of glycosylation type Ic (CDG‐Ic). Their clinical presentation was mainly neurological with developmental retardation, muscular hypotonia, and epilepsy. Several symptoms commonly seen in CDG‐Ia such as inverted nipples, abnormal fat distribution, and cerebellar hypoplasia were not observed. The clinical course is milder overall, with a better neurological outcome, than in CDG‐Ia. The isoelectric focusing pattern of serum transferrin in CDG‐Ia and CDG‐Ic is indistinguishable. Interestingly, β‐trace protein in cerebrospinal fluid derived from immunoblot analysis of the brain showed a less pronounced hypoglycosylation pattern in CDG‐Ic patients than in CDG‐Ia patients. Analysis of lipid‐linked oligosaccharides revealed an accumulation of Man9GlcNAc2 intermediates due to dolichol pyrophosphate–Man9GlcNAc2 α‐1,3 glucosyltransferase deficiency. All patients were homozygous for an A333V mutation. Ann Neurol 2000;47:776–781

Improvement of dolichol-linked oligosaccharide biosynthesis by the squalene synthase inhibitor zaragozic acid.

The majority of congenital disorders of glycosylation (CDG) are caused by defects of dolichol (Dol)-linked oligosaccharide assembly, which lead to under-occupancy of N-glycosylation sites. Most mutations encountered in CDG are hypomorphic, thus leaving residual activity to the affected biosynthetic enzymes. We hypothesized that increased cellular levels of Dol-linked substrates might compensate for the low biosynthetic activity and thereby improve the output of protein N-glycosylation in CDG. To this end, we investigated the potential of the squalene synthase inhibitor zaragozic acid A to redirect the flow of the polyisoprene pathway toward Dol by lowering cholesterol biosynthesis. The addition of zaragozic acid A to CDG fibroblasts with a Dol-P-Man synthase defect led to the formation of longer Dol-P species and to increased Dol-P-Man levels. This treatment was shown to decrease the pathologic accumulation of incomplete Dol pyrophosphate-GlcNAc2Man5 in Dol-P-Man synthase-deficient fibroblasts. Zaragozic acid A treatment also decreased the amount of truncated protein N-linked oligosaccharides in these CDG fibroblasts. The increased cellular levels of Dol-P-Man and possibly the decreased cholesterol levels in zaragozic acid A-treated cells also led to increased availability of the glycosylphosphatidylinositol anchor as shown by the elevated cell-surface expression of the CD59 protein. This study shows that manipulation of the cellular Dol pool, as achieved by zaragozic acid A addition, may represent a valuable approach to improve N-linked glycosylation in CDG cells.