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Congenital disorder of glycosylation type Ig (ALG12‐CDG) is a rare inherited metabolic disease caused by a defect in alpha‐mannosyltransferase 8, encoded by the ALG12 gene (22q13.33). To date, only 15 patients have been diagnosed with ALG12‐CDG globally. Due to a newborn Slovak patient's clinical and biochemical abnormalities, the isoelectric focusing of transferrin was performed with observed significant hypoglycosylation typical of CDG I. Furthermore, analysis of neutral serum N‐glycans by mass spectrometry revealed the accumulation of GlcNAc2Man5–7 and decreased levels of GlcNAc2Man8–9, which indicated impaired ALG12 enzymatic activity. Genetic analysis of the coding regions of the ALG12 gene of the patient revealed a novel homozygous substitution mutation c.1439T>C p.(Leu480Pro) within Exon 10. Furthermore, both of the patient's parents and his twin sister were asymptomatic heterozygous carriers of the variant. This comprehensive genomic and glycomic approach led to the confirmation of the ALG12 pathogenic variant responsible for the clinical manifestation of the disorder in the patient described. [ABSTRACT FROM AUTHOR]

Abstract Background Congenital disorder of glycosylation (CDG) type I is a group of rare disorders caused by recessive mutations in up to 25 genes that impair the N‐glycan precursor formation and its transfer to proteins resulting in hypoglycosylation of multiple proteins. Congenital disorder of glycosylation causes multisystem defects usually with psychomotor delay that is diagnosed in the infancy. We aim to supply further evidences supporting that CDG may be underestimated. Methods Antithrombin and factor XI were studied by chromogenic and coagulometric methods. Hypoglycosylation of plasma proteins was evaluated by western blot, HPLC, Q‐TOF, and RP‐LC‐MRM‐MS. Genetic analysis included whole exome, Sanger sequencing, and PCR‐allele specific assay. Results We here present an intriguing patient with an exceptional phenotype: 25‐year‐old women with a ventricular septal defect and severe idiopathic scoliosis but no facial dysmorphism, who dances as a professional, and has a University degree. Congenital disorder of glycosylation diagnosis started through the identification of antithrombin deficiency without SERPINC1 defect and the detection of hypoglycosylated forms. Increased levels of hypoglycosylated forms of F XI (also with significant deficiency) and transferrin were also detected. Whole exome analysis showed a novel homozygous ALG12 variant c.77T>A, p.(Val26Asp) supporting an ALG12‐CDG diagnosis. It also showed three new variants in KMT2D, and a mild, known ALG6 variant. Conclusions This novel ALG12‐CDG patient (the 13th reported) underlines the heterogeneity of this CDG and broadens its phenotypical spectrum, supports that these disorders are underestimated, and suggests that combination of global hypoglycosylation with specific gene defects might determine the clinical manifestations of CDG patients.

Background: Congenital disorder of glycosylation (CDG) type I is a group of rare disorders caused by recessive mutations in up to 25 genes that impair the N‐glycan precursor formation and its transfer to proteins resulting in hypoglycosylation of multiple proteins. Congenital disorder of glycosylation causes multisystem defects usually with psychomotor delay that is diagnosed in the infancy. We aim to supply further evidences supporting that CDG may be underestimated. Methods: Antithrombin and factor XI were studied by chromogenic and coagulometric methods. Hypoglycosylation of plasma proteins was evaluated by western blot, HPLC, Q‐TOF, and RP‐LC‐MRM‐MS. Genetic analysis included whole exome, Sanger sequencing, and PCR‐allele specific assay. Results: We here present an intriguing patient with an exceptional phenotype: 25‐year‐old women with a ventricular septal defect and severe idiopathic scoliosis but no facial dysmorphism, who dances as a professional, and has a University degree. Congenital disorder of glycosylation diagnosis started through the identification of antithrombin deficiency without SERPINC1 defect and the detection of hypoglycosylated forms. Increased levels of hypoglycosylated forms of F XI (also with significant deficiency) and transferrin were also detected. Whole exome analysis showed a novel homozygous ALG12 variant c.77T>A, p.(Val26Asp) supporting an ALG12‐CDG diagnosis. It also showed three new variants in KMT2D, and a mild, known ALG6 variant. Conclusions: This novel ALG12‐CDG patient (the 13th reported) underlines the heterogeneity of this CDG and broadens its phenotypical spectrum, supports that these disorders are underestimated, and suggests that combination of global hypoglycosylation with specific gene defects might determine the clinical manifestations of CDG patients. [ABSTRACT FROM AUTHOR]

ALG12-congenital disorder of glycosylation (ALG12-CDG) is a rare disorder caused by a deficiency of dolichol-P-mannose:Man 7 GlcNAc 2 -PP-dolichyl-α-6-mannosyltransferase which presents with intellectual disability, hypotonia, dysmorphic features, low IgG levels with recurrent infections, male genital hypoplasia, and coagulation abnormalities. We report a unique family with three affected individuals, including two older brothers with only cognitive and coagulation defects and a younger brother who died from a severe multisystem disease at age 18 months. The two living brothers are the oldest and mildest cases of ALG12-CDG described thus far. Whole exome sequencing of the older brothers revealed a previously described c.1001delA (p.N334TfsX15) pathogenic variant and a c.671C > T (p.T224 M) variant of uncertain significance in ALG12. Our cases broaden the recognized genetic and phenotypic spectrum of this disorder and suggest a role for other genetic and environmental factors in modulating disease phenotype. [ABSTRACT FROM AUTHOR]

Congenital disorders of glycosylation (CDG) are genetic diseases characterized by deficient synthesis (CDG type I) and/or abnormal processing (CDG type II) of glycan moieties linked to protein and lipids. The impact of the molecular defects on protein glycosylation and in turn on the clinical phenotypes of patients with CDG is not yet understood. ALG12-CDG is due to deficiency of ALG12 α1,6-mannosyltransferase that adds the eighth mannose residue on the dolichol-PP-oligosaccharide precursor in the endoplasmic reticulum. ALG12-CDG is a severe multisystem disease associated with low to deficient serum immunoglobulins and recurrent infections. We thoroughly investigated the glycophenotype in a patient with novel ALG12 variants and immunodeficiency. We analyzed serum native transferrin, as first line test for CDG and we profiled serum IgG and total serum N-glycans by a combination of consolidated (N-glycan analysis by MALDI MS) and innovative mass spectrometry-based protocols, such as GlycoWorks RapiFluor N-glycan analysis coupled with LC-ESI MS. Intact serum transferrin showed, as expected for a CDG type I defect, underoccupancy of N-glycosylation sites. Surprisingly, total serum proteins and IgG N-glycans showed some specific changes, consisting in accumulating amounts of definite high-mannose and hybrid structures. As a whole, ALG12-CDG behaves as a dual CDG (CDG-I and II defects) and it is associated with distinct, abnormal glycosylation of total serum and IgG N-glycans. Glycan profiling of target glycoproteins may endorse the molecular defect unraveling the complex clinical phenotype of CDG patients. [ABSTRACT FROM AUTHOR]

We reported an oligosaccharide diphosphodolichol (DLO) diphosphatase (DLODP) that generates dolichyl-phosphate and oligosaccharyl phosphates (OSPs) from DLO in vitro. This enzyme could underlie cytoplasmic OSP generation and promote dolichyl-phosphate recycling from truncated endoplasmic reticulum (ER)-generated DLO intermediates. However, during subcellular fractionation, DLODP distribution is closer to that of a Golgi apparatus (GA) marker than those of ER markers. Here, we examined the effect of brefeldin A (BFA), which fuses the GA with the ER on OSP metabolism. In order to increase the steady state level of truncated DLO while allowing formation of mature DLO (Glc3Man9GlcNAc2-PP-dolichol), dolichyl-P-mannose Man7GlcNAc2-PP-dolichol mannosyltransferase was partially downregulated in HepG2 cells. We show that BFA provokes GA endomannosidase trimming of Glc3Man9GlcNAc2-PP-dolichol to yield a Man8GlcNAc2-PP-dolichol structure that does not give rise to cytoplasmic Man8GlcNAc2-P. BFA also strikingly increased OSP derived from mature DLO within the endomembrane system without affecting levels of Man7GlcNAc2-PP-dolichol or cytoplasmic Man7GlcNAc2-P. The BFA-provoked increase in endomembrane-situated OSP is sensitive to nocodazole, and BFA causes partial redistribution of DLODP activity from GA- to ER-containing regions of density gradients. These findings are consistent with BFA-provoked microtubule-dependent GA-to-ER transport of a previously reported DLODP that acts to generate a novel endomembrane-situated OSP population.

Congenital Disorder of Glycosylation type Ig (ALG12-CDG) is part of a group of autosomal recessive conditions caused by deficiency of proteins involved in the assembly of dolichol-oligosaccharides used for protein N-glycosylation. In ALG12-CDG, the enzyme affected is encoded by the ALG12 gene. Affected individuals present clinically with neurodevelopmental delay, growth retardation, immune deficiency, male genital hypoplasia, and cardiomyopathy. A total of six individuals have been reported in the literature. Here, we present an infant with rhizomelic short stature, talipes equinovarus, platyspondyly, and joint dislocations. The infant had marked under-ossification of the pubic bones. Exome sequencing was performed and two deletions, each resulting in a frameshift, were found in ALG12. A review of the literature revealed two infants with ALG12-CDG and a severe skeletal dysplasia, including under-ossification of cervical vertebrae, pubic bones, and knees; in addition to talipes equinovarus and rhizomelic short stature. The phenotype of the individual we describe resembles pseudodiastrophic dysplasia and we discuss similarities and differences between ALG12-CDG and pseudodiastrophic dysplasia. The differential diagnosis in selected undiagnosed skeletal dysplasias should include CDGs.

Background: Congenital disorder of glycosylation (CDG) type I is a group of rare disorders caused by recessive mutations in up to 25 genes that impair the N-glycan precursor formation and its transfer to proteins resulting in hypoglycosylation of multiple proteins. Congenital disorder of glycosylation causes multisystem defects usually with psychomotor delay that is diagnosed in the infancy. We aim to supply further evidences supporting that CDG may be underestimated., Methods: Antithrombin and factor XI were studied by chromogenic and coagulometric methods. Hypoglycosylation of plasma proteins was evaluated by western blot, HPLC, Q-TOF, and RP-LC-MRM-MS. Genetic analysis included whole exome, Sanger sequencing, and PCR-allele specific assay., Results: We here present an intriguing patient with an exceptional phenotype: 25-year-old women with a ventricular septal defect and severe idiopathic scoliosis but no facial dysmorphism, who dances as a professional, and has a University degree. Congenital disorder of glycosylation diagnosis started through the identification of antithrombin deficiency without SERPINC1 defect and the detection of hypoglycosylated forms. Increased levels of hypoglycosylated forms of F XI (also with significant deficiency) and transferrin were also detected. Whole exome analysis showed a novel homozygous ALG12 variant c.77T>A, p.(Val26Asp) supporting an ALG12-CDG diagnosis. It also showed three new variants in KMT2D, and a mild, known ALG6 variant., Conclusions: This novel ALG12-CDG patient (the 13th reported) underlines the heterogeneity of this CDG and broadens its phenotypical spectrum, supports that these disorders are underestimated, and suggests that combination of global hypoglycosylation with specific gene defects might determine the clinical manifestations of CDG patients., (© 2020 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals LLC.)

We reported an oligosaccharide diphosphodolichol (DLO) diphosphatase (DLODP) that generates dolichyl-phosphate and oligosaccharyl phosphates (OSPs) from DLO in vitro. This enzyme could underlie cytoplasmic OSP generation and promote dolichyl-phosphate recycling from truncated endoplasmic reticulum (ER)-generated DLO intermediates. However, during subcellular fractionation, DLODP distribution is closer to that of a Golgi apparatus (GA) marker than those of ER markers. Here, we examined the effect of brefeldin A (BFA), which fuses the GA with the ER on OSP metabolism. In order to increase the steady state level of truncated DLO while allowing formation of mature DLO (Glc3Man9GlcNAc2-PP-dolichol), dolichyl-P-mannose Man7GlcNAc2-PP-dolichol mannosyltransferase was partially downregulated in HepG2 cells. We show that BFA provokes GA endomannosidase trimming of Glc3Man9GlcNAc2-PP-dolichol to yield a Man8GlcNAc2-PP-dolichol structure that does not give rise to cytoplasmic Man8GlcNAc2-P. BFA also strikingly increased OSP derived from mature DLO within the endomembrane system without affecting levels of Man7GlcNAc2-PP-dolichol or cytoplasmic Man7GlcNAc2-P. The BFA-provoked increase in endomembrane-situated OSP is sensitive to nocodazole, and BFA causes partial redistribution of DLODP activity from GA- to ER-containing regions of density gradients. These findings are consistent with BFA-provoked microtubule-dependent GA-to-ER transport of a previously reported DLODP that acts to generate a novel endomembrane-situated OSP population., (Copyright © 2016 by the American Society for Biochemistry and Molecular Biology, Inc.)

Congenital Disorder of Glycosylation type Ig (ALG12-CDG) is part of a group of autosomal recessive conditions caused by deficiency of proteins involved in the assembly of dolichol-oligosaccharides used for protein N-glycosylation. In ALG12-CDG, the enzyme affected is encoded by the ALG12 gene. Affected individuals present clinically with neurodevelopmental delay, growth retardation, immune deficiency, male genital hypoplasia, and cardiomyopathy. A total of six individuals have been reported in the literature. Here, we present an infant with rhizomelic short stature, talipes equinovarus, platyspondyly, and joint dislocations. The infant had marked underossification of the pubic bones. Exome sequencing was performed and two deletions, each resulting in frameshifts, were found in ALG12 . A review of the literature revealed two infants with ALG12-CDG and a severe skeletal dysplasia, including under-ossification of cervical vertebrae, pubic bones, and knees; in addition to talipes equinovarus and rhizomelic short stature. The phenotype of the individual we describe resembles pseudodiastrophic dysplasia and we discuss similarities and differences between ALG12-CDG and pseudodiastrophic dysplasia. The differential diagnosis in selected undiagnosed skeletal dysplasias should include CDGs.