Multiple phenotypes in phosphoglucomutase 1 deficiency.

BACKGROUND: Congenital disorders of glycosylation are genetic syndromes that result in impaired glycoprotein production. We evaluated patients who had a novel recessive disorder of glycosylation, with a range of clinical manifestations that included hepatopathy, bifid uvula, malignant hyperthermia, hypogonadotropic hypogonadism, growth retardation, hypoglycemia, myopathy, dilated cardiomyopathy, and cardiac arrest. METHODS: Homozygosity mapping followed by whole-exome sequencing was used to identify a mutation in the gene for phosphoglucomutase 1 (PGM1) in two siblings. Sequencing identified additional mutations in 15 other families. Phosphoglucomutase 1 enzyme activity was assayed on cell extracts. Analyses of glycosylation efficiency and quantitative studies of sugar metabolites were performed. Galactose supplementation in fibroblast cultures and dietary supplementation in the patients were studied to determine the effect on glycosylation. RESULTS: Phosphoglucomutase 1 enzyme activity was markedly diminished in all patients. Mass spectrometry of transferrin showed a loss of complete N-glycans and the presence of truncated glycans lacking galactose. Fibroblasts supplemented with galactose showed restoration of protein glycosylation and no evidence of glycogen accumulation. Dietary supplementation with galactose in six patients resulted in changes suggestive of clinical improvement. A new screening test showed good discrimination between patients and controls. CONCLUSIONS: Phosphoglucomutase 1 deficiency, previously identified as a glycogenosis, is also a congenital disorder of glycosylation. Supplementation with galactose leads to biochemical improvement in indexes of glycosylation in cells and patients, and supplementation with complex carbohydrates stabilizes blood glucose. A new screening test has been developed but has not yet been validated. (Funded by the Netherlands Organization for Scientific Research and others.).

Perinatal and early infantile symptoms in congenital disorders of glycosylation.

Congenital disorders of glycosylation (CDG) are a rapidly growing family of inborn errors. Screening for CDG in suspected cases is usually performed in the first year of life by serum transferrin isoelectric focusing or mass spectrometry. Based on the transferrin analysis patients can be biochemically diagnosed with a type 1 or type 2 transferrin pattern, and labeled as CDG-I, or CDG-II. The diagnosis of CDG is frequently delayed due to the highly variable phenotype, some cases showing single organ involvement and others mimicking syndromes, like skeletal dysplasia, cutis laxa syndrome, or congenital muscle dystrophy. The aim of our study was to evaluate perinatal abnormalities and early discriminative symptoms in 58 patients consecutively diagnosed with diverse CDG-subtypes. Neonatal findings and clinical features in the first months of life were studied in 36 children with CDG-I and 22 with CDG-II. Maternal complications were found in five, small for gestational age in nine patients. Five children had abnormal neonatal screening results for hypothyroidism. Congenital microcephaly and neonatal seizures were common in CDG-II. Inverted nipples were uncommon with 5 out of 58 children. Dysmorphic features were mostly nonspecific, except for cutis laxa. Early complications included feeding problems, cardiomyopathy, thrombosis, and bleeding. Cases presenting in the neonatal period had the highest mortality rate. Survival in CDG patients is highly dependent on early intervention therapy. We recommend low threshold screening for glycosylation disorders in infants with neurologic symptoms, even in the absence of abnormal fat distribution. Growth retardation and neonatal bleeding increase suspicion for CDG.

A novel cerebello-ocular syndrome with abnormal glycosylation due to abnormalities in dolichol metabolism.

Cerebellar hypoplasia and slowly progressive ophthalmological symptoms are common features in patients with congenital disorders of glycosylation type I. In a group of patients with congenital disorders of glycosylation type I with unknown aetiology, we have previously described a distinct phenotype with severe, early visual impairment and variable eye malformations, including optic nerve hypoplasia, retinal coloboma, congenital cataract and glaucoma. Some of the symptoms overlapped with the phenotype in other congenital disorders of glycosylation type I subtypes, such as vermis hypoplasia, anaemia, ichtyosiform dermatitis, liver dysfunction and coagulation abnormalities. We recently identified pathogenic mutations in the SRD5A3 gene, encoding steroid 5α-reductase type 3, in a group of patients who presented with this particular phenotype and a common metabolic pattern. Here, we report on the clinical, genetic and metabolic features of 12 patients from nine families with cerebellar ataxia and congenital eye malformations diagnosed with SRD5A3-congenital disorders of glycosylation due to steroid 5α-reductase type 3 defect. This enzyme is necessary for the reduction of polyprenol to dolichol, the lipid anchor for N-glycosylation in the endoplasmic reticulum. Dolichol synthesis is an essential metabolic step in protein glycosylation. The current defect leads to a severely abnormal glycosylation state already in the early phase of the N-glycan biosynthesis pathway in the endoplasmic reticulum. We detected high expression of SRD5A3 in foetal brain tissue, especially in the cerebellum, consistent with the finding of the congenital cerebellar malformations. Based on the overlapping clinical, biochemical and genetic data in this large group of patients with congenital disorders of glycosylation, we define a novel syndrome of cerebellar ataxia associated with congenital eye malformations due to a defect in dolichol metabolism.

SRD5A3 is required for converting polyprenol to dolichol and is mutated in a congenital glycosylation disorder.

N-linked glycosylation is the most frequent modification of secreted and membrane-bound proteins in eukaryotic cells, disruption of which is the basis of the congenital disorders of glycosylation (CDGs). We describe a new type of CDG caused by mutations in the steroid 5alpha-reductase type 3 (SRD5A3) gene. Patients have mental retardation and ophthalmologic and cerebellar defects. We found that SRD5A3 is necessary for the reduction of the alpha-isoprene unit of polyprenols to form dolichols, required for synthesis of dolichol-linked monosaccharides, and the oligosaccharide precursor used for N-glycosylation. The presence of residual dolichol in cells depleted for this enzyme suggests the existence of an unexpected alternative pathway for dolichol de novo biosynthesis. Our results thus suggest that SRD5A3 is likely to be the long-sought polyprenol reductase and reveal the genetic basis of one of the earliest steps in protein N-linked glycosylation.

Loss-of-function mutations in ATP6V0A2 impair vesicular trafficking, tropoelastin secretion and cell survival.

Autosomal recessive cutis laxa type 2 (ARCL2), a syndrome of growth and developmental delay and redundant, inelastic skin, is caused by mutations in the a2 subunit of the vesicular ATPase H+-pump (ATP6V0A2). The goal of this study was to define the disease mechanisms that lead to connective tissue lesions in ARCL2. In a new cohort of 17 patients, DNA sequencing of ATP6V0A2 detected either homozygous or compound heterozygous mutations. Considerable allelic and phenotypic heterogeneity was observed, with a missense mutation of a moderately conserved residue p.P87L leading to unusually mild disease. Abnormal N- and/or mucin type O-glycosylation was observed in all patients tested. Premature stop codon mutations led to decreased ATP6V0A2 mRNA levels by destabilizing the mutant mRNA via the nonsense-mediated decay pathway. Loss of ATP6V0A2 either by siRNA knockdown or in ARCL2 cells resulted in distended Golgi cisternae, accumulation of abnormal lysosomes and multivesicular bodies. Immunostaining of ARCL2 cells showed the accumulation of tropoelastin (TE) in the Golgi and in large, abnormal intracellular and extracellular aggregates. Pulse-chase studies confirmed impaired secretion and increased intracellular retention of TE, and insoluble elastin assays showed significantly reduced extracellular deposition of mature elastin. Fibrillin-1 microfibril assembly and secreted lysyl oxidase activity were normal in ARCL2 cells. TUNEL staining demonstrated increased rates of apoptosis in ARCL2 cell cultures. We conclude that loss-of-function mutations in ATP6V0A2 lead to TE aggregation in the Golgi, impaired clearance of TE aggregates and increased apoptosis of elastogenic cells.