DDOST‐CDG: Clinical and molecular characterization of a third patient with a milder and a predominantly movement disorder phenotype

Abstract

Congenital disorders of glycosylation (CDG) are a group of heterogeneous inherited metabolic disorders affecting posttranslational protein modification. DDOST-CDG, caused by biallelic pathogenic variants in DDOST which encodes dolichyl-diphospho-oligosaccharide-protein glycosyltransferase, a subunit of N-glycosylation oligosaccharyltransferase (OST) complex, is an ultrarare condition that has been described in two patients only. The main clinical features in the two reported patients include profound developmental delay, failure to thrive, and hypotonia. In addition, both patients had abnormal transferrin glycosylation. Here, we report an 18-year-old male who presented with moderate developmental delay, progressive opsoclonus, myoclonus, ataxia, tremor, and dystonia. Biochemical studies by carbohydrate deficient transferrin analysis showed a type I CDG pattern. Exome sequencing identified compound heterozygous variants in DDOST: a maternally inherited variant, c.1142dupT (p.Leu381Phefs*11), and a paternally inherited variant, c.661 T > C (p.Ser221Pro). Plasma N-glycan profiling showed mildly increased small high mannose glycans including Man 0-5 GlcNAc 2, a pattern consistent with what was previously reported in DDOST-CDG or defects in other subunits of OST complex. Western blot analysis on patient's fibroblasts revealed decreased expression of DDOST and reduced intracellular N-glycosylation, as evident by the biomarkers ICAM-1 and LAMP2. Our study highlights the clinical variability, expands the clinical and biochemical phenotypes, and describes new genotype, which all are essential for diagnosing and managing patients with DDOST-CDG.

Functional classification of DDOST variants of uncertain clinical significance in congenital disorders of glycosylation

“…Since OST48 is essential for LOX N -glycosylation, we tested if we could use this in a functional complementation assay to discriminate benign from pathogenic DDOST VUS. As proof of principle, we complemented DDOST -depleted U87 cells with wild-type DDOST cDNA and five mutations reported in 3 DDOST-CDG patients 13 , 41 , 42 , and analysed the impact on LOX N -glycosylation under hypoxic conditions. We used CRISPR/Cas9 to target the DDOST splice donor (sg DDOST -sd) site at exon 2 to disrupt endogenous DDOST and in parallel complemented the cells with a DDOST cDNA that is intrinsically resistant to this sgRNA.…”

“…Parental cells are shown for reference. Cells were re-seeded 7 days after transduction and moved to hypoxia (1% O 2 ) on day 8 for 24 h. Variants in red were reported previously 13 , 41 , 42 . Data are plotted mean ± s.d.…”

“…Analysis of large cohorts of affected individuals may reveal genotype–phenotype correlations, but because patient numbers are small and there is enormous genetic heterogeneity, correlations such as these are difficult to establish for CDG. Hypoglycosylation has been confirmed in patient-derived fibroblasts to validate a small number of DDOST variants as pathogenic 13 , 42 , but complementation with wild-type DDOST cDNA in patient-derived fibroblasts is laborious and challenging and does not lend itself to a robust and effective basis for an assay 13 . Moreover, this approach does not allow testing of individual VUS in patients with different mutations on the two DDOST alleles.…”

Single-Cell Transcriptome Analysis Reveals Development-Specific Networks at Distinct Synchronized Antral Follicle Sizes in Sheep Oocytes

“…In addition, we identified 15 co-expressed signaling pathways in the LO group (Figure 4B), and, among these, the biosynthesis of amino acids and various types of N-glycan, arginine, and proline metabolism, and protein processing in the endoplasmic reticulum play a key role in oocyte maturation. Due to the critical role of glycans in protein modification and the regulation of diverse cellular functions [34], we further analyzed the N-glycan biosynthesis pathway and found 12 highly expressed genes, including MGAT4B [35], ALG3 [36], and DDOST [37], suggesting that N-glycosylation was one of the key factors affecting sheep oocyte maturation.…”