Identification and characterization of a missense mutation in the O-linked β-N-acetylglucosamine (O-GlcNAc) transferase gene that segregates with X-linked intellectual disability

Abstract

-GlcNAc is a regulatory post-translational modification of nucleocytoplasmic proteins that has been implicated in multiple biological processes, including transcription. In humans, single genes encode enzymes for its attachment (-GlcNAc transferase (OGT)) and removal (-GlcNAcase (OGA)). An X-chromosome exome screen identified a missense mutation, which encodes an amino acid in the tetratricopeptide repeat, in (759G>T (p.L254F)) that segregates with X-linked intellectual disability (XLID) in an affected family. A decrease in steady-state OGT protein levels was observed in isolated lymphoblastoid cell lines from affected individuals, consistent with molecular modeling experiments. Recombinant expression of L254F-OGT demonstrated that the enzyme is active as both a glycosyltransferase and an HCF-1 protease. Despite the reduction in OGT levels seen in the L254F-OGT individual cells, we observed that steady-state global-GlcNAc levels remained grossly unaltered. Surprisingly, lymphoblastoids from affected individuals displayed a marked decrease in steady-state OGA protein and mRNA levels. We observed an enrichment of the OGT-containing transcriptional repressor complex mSin3A-HDAC1 at the proximal promoter region of and correspondingly decreased promoter activity in affected cells. Global transcriptome analysis of L254F-OGT lymphoblastoids compared with controls revealed a small subset of genes that are differentially expressed. Thus, we have begun to unravel the molecular consequences of the 759G>T (p.L254F) mutation in that uncovered a compensation mechanism, albeit imperfect, given the phenotype of affected individuals, to maintain steady-state-GlcNAc levels. Thus, a single amino acid substitution in the regulatory domain (the tetratricopeptide repeat domain) of OGT, which catalyzes the -GlcNAc post-translational modification of nuclear and cytosolic proteins, appears causal for XLID.

A missense mutation in the catalytic domain of O‐GlcNAc transferase links perturbations in protein O‐GlcNAcylation to X‐linked intellectual disability

“…B). This hints at the existence of a compensatory mechanism that allows cells to partially maintain O ‐GlcNAc homeostasis by reducing OGA levels—as also observed in some of the other OGT XLID mutations in the TPR domain . Thus, the N648Y mutation leads to hypoglycosylation in mES cells.…”

Roles of the HUWE1 ubiquitin ligase in nervous system development, function and disease

“…To date, 7 different mutations in OGT have been found in 11 individuals with ID. Familial inheritance indicates that OGT mutations segregate with ID, providing further support for the involvement of OGT in ID [96][97][98].…”

“…OGT was found to function with EEL-1 to regulate inhibitory GABA neuron function. Similar to HUWE1, human genetic studies have identified OGT mutations in multiple families with non-syndromic ID [80,[95][96][97][98]. To date, 7 different mutations in OGT have been found in 11 individuals with ID.…”

Chemical and Biochemical Strategies To Explore the Substrate Recognition of O‐GlcNAc‐Cycling Enzymes

“…The TPR region folds into a super α‐helix and has been suggested to play an important role in protein interactions . Furthermore, mutations in the TPR region have been implicated in intellectual disability . Three isoforms of human OGT have been reported, which differ only in the number of TPR repeats: 13.5 for ncOGT (the most abundant isoform in the nucleus and cytoplasm), nine for mOGT (mitochondria), and 2.5 for a short isoform, sOGT.…”

“…[27,28] Furthermore, mutationsi nt he TPR region have been implicated in intellectual disability. [29][30][31] Three isoformso fh uman OGT have been reported, [32,33] which differ only in the number of TPRr epeats:1 3.5 for ncOGT( the most abundant isoform in the nucleusand cytoplasm), nine for mOGT (mitochondria), and 2.5 for as hort isoform, sOGT.H owever,t he physiological importance of the two shorter isoforms remain mostly unknown. [34][35][36] Several studies have reported that the extended TPR region is necessary for binding and glycosylating protein-but not peptide-substrates.…”