Dataset Information

Mutations in STT3A and STT3B cause two congenital disorders of glycosylation.

ABSTRACT: We describe two unreported types of congenital disorders of glycosylation (CDG) which are caused by mutations in different isoforms of the catalytic subunit of the oligosaccharyltransferase (OST). Each isoform is encoded by a different gene (STT3A or STT3B), resides in a different OST complex and has distinct donor and acceptor substrate specificities with partially overlapping functions in N-glycosylation. The two cases from unrelated consanguineous families both show neurologic abnormalities, hypotonia, intellectual disability, failure to thrive and feeding problems. A homozygous mutation (c.1877T > C) in STT3A causes a p.Val626Ala change and a homozygous intronic mutation (c.1539 + 20G > T) in STT3B causes the other disorder. Both mutations impair glycosylation of a GFP biomarker and are rescued with the corresponding cDNA. Glycosylation of STT3A- and STT3B-specific acceptors is decreased in fibroblasts carrying the corresponding mutated gene and expression of the STT3A (p.Val626Ala) allele in STT3A-deficient HeLa cells does not rescue glycosylation. No additional cases were found in our collection or in reviewing various databases. The STT3A mutation significantly impairs glycosylation of the biomarker transferrin, but the STT3B mutation only slightly affects its glycosylation. Additional cases of STT3B-CDG may be missed by transferrin analysis and will require exome or genome sequencing.

SUBMITTER: Shrimal S

PROVIDER: S-EPMC3888133 | biostudies-literature | 2013 Nov

REPOSITORIES: biostudies-literature

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Mutations in STT3A and STT3B cause two congenital disorders of glycosylation.

Shrimal Shiteshu S Ng Bobby G BG Losfeld Marie-Estelle ME Gilmore Reid R Freeze Hudson H HH

Human molecular genetics 20130710 22

We describe two unreported types of congenital disorders of glycosylation (CDG) which are caused by mutations in different isoforms of the catalytic subunit of the oligosaccharyltransferase (OST). Each isoform is encoded by a different gene (STT3A or STT3B), resides in a different OST complex and has distinct donor and acceptor substrate specificities with partially overlapping functions in N-glycosylation. The two cases from unrelated consanguineous families both show neurologic abnormalities, ...[more]

PMID: 23842455

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Project description:STT3A and STT3B are the main catalytic subunit of oligosaccharyltransferase (OST-A and OST-B in mammalian cells), which primarily mediate cotranslational and posttranslocational N-linked glycosylation, respectively. To determine the specificity of STT3A and STT3B, we performed proteomics and glycoproteomics analyses to characterize the protein expression and glycosylation in STT3A-knockout (KO), STT3B-KO, and wild-type (WT) HEK293 cells. In total, 3,993 proteins, 4,020 unique N-linked intact glycopeptides (IGPs) and 723 glycosites, representing 401 glycoproteins were identified in KO and WT cells. Deletion of the STT3A gene had a greater impact on the protein expression than deletion of STT3B, especially on glycoproteins. In addition, mannosylated N-glycans from glycoproteins were reduced, while fucosylated N-glycans were increased in STT3A-KO cells. The glycan changes may be caused by the differential expression of glycosyltransferases and the activation of unfolded protein response (UPR) with further analysis of glycan-related enzymes. Interestingly, hyperglycosylated proteins were identified in KO cells. We verified that the hyperglycosylation of ENPL was caused by the ER stress and UPR, which were induced by the deletion of the STT3A. Overall, the specificity of STT3A and STT3B revealed that defects in the OST subunit not only broadly affect N-linked glycosylation of the protein but also affect protein expression.STT3A and STT3B are the main catalytic subunit of oligosaccharyltransferase (OST-A and OST-B in mammalian cells), which primarily mediate cotranslational and posttranslocational N-linked glycosylation, respectively. To determine the specificity of STT3A and STT3B, we performed proteomics and glycoproteomics analyses to characterize the protein expression and glycosylation in STT3A-knockout (KO), STT3B-KO, and wild-type (WT) HEK293 cells. In total, 3,993 proteins, 4,020 unique N-linked intact glycopeptides (IGPs) and 723 glycosites, representing 401 glycoproteins were identified in KO and WT cells. Deletion of the STT3A gene had a greater impact on the protein expression than deletion of STT3B, especially on glycoproteins. In addition, mannosylated N-glycans from glycoproteins were reduced, while fucosylated N-glycans were increased in STT3A-KO cells. The glycan changes may be caused by the differential expression of glycosyltransferases and the activation of unfolded protein response (UPR) with further analysis of glycan-related enzymes. Interestingly, hyperglycosylated proteins were identified in KO cells. We verified that the hyperglycosylation of ENPL was caused by the ER stress and UPR, which were induced by the deletion of the STT3A. Overall, the specificity of STT3A and STT3B revealed that defects in the OST subunit not only broadly affect N-linked glycosylation of the protein but also affect protein expression.

Project description:The application of genome sequencing in patients with intellectual disability and congenital anomalies (ID/CA) typically identifies causative mutations only in a minority of cases. We hypothesized that some cases of ID/CA are caused by epigenetic aberrations that dysregulate normal genome function, and that these would be missed by conventional sequencing approaches. We performed Illumina 450k DNA methylation profiling in ~500 individuals with ID/CA, who were negative for causative mutations by microarray, exome- and/or whole genome sequencing. We screened patient methylation profiles for epimutations absent in ~1,500 controls, and validated these by bisulfite sequencing, revealing that epimutations represent large methylation changes specifically on one allele. Parental studies show that approximately half represent de novo events, a rate significantly higher than that seen in controls. We identified seven recurrent events, two of which (FMR1, MEG3) have known disease associations, validating our method for detecting pathogenic epimutations. From large-scale sequencing, population and expression studies we conclude that epimutations are: (i) Frequently associated with extreme outlier and mono-allelic gene expression, with an impact comparable to loss-of-function mutations; (ii) Generally conserved across multiple tissues within an individual, validating the use of blood DNA to study ID/CA; (iii) Can occur secondary to cis-linked regulatory mutations, providing a rationale for interpreting non-coding genetic variants; (iv) Occur sporadically with a remarkably high de novo rate, and (v) Exhibit non-Mendelian inheritance, likely being reset between generations by epigenetic reprogramming during embryogenesis. We conclude that epimutations underlie 5-10% of patients with ID/CA who are refractory to conventional mutation screening, and propose that epigenome profiling represents a promising method for the study of human disease that complements sequence-based approaches.

Dataset Information

Mutations in STT3A and STT3B cause two congenital disorders of glycosylation.

Publications

Mutations in STT3A and STT3B cause two congenital disorders of glycosylation.

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