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Glycobiology Mar 2010The bifunctional enzyme UDP-GlcNAc 2-epimerase/ ManNAc kinase (GNE/MNK), encoded by the GNE gene, catalyzes the first two committed, rate-limiting steps in the...
The bifunctional enzyme UDP-GlcNAc 2-epimerase/ ManNAc kinase (GNE/MNK), encoded by the GNE gene, catalyzes the first two committed, rate-limiting steps in the biosynthesis of N-acetylneuraminic acid (sialic acid). GNE/MNK is feedback inhibited by binding of the downstream product, CMP-sialic acid in its allosteric site. GNE mutations can result in two human disorders, hereditary inclusion body myopathy (HIBM) or sialuria. So far, no active site geometry predictions or conformational transitions involved with function are available for mammalian GNE/MNK. The N-terminal GNE domain is homologous to various prokaryotic 2-epimerases, some of which have solved crystallographic structures. The C-terminal MNK domain belongs to the sugar kinases superfamily; its crystallographic structure is solved at 2.84 A and three-dimensional structures have also been reported for several other kinases. In this work, we employed available structural data of GNE/MNK homologs to model the active sites of human GNE/MNK and identify critical amino acid residues responsible for interactions with substrates. In addition, we modeled effects of GNE/MNK missense mutations associated with HIBM or sialuria on helix arrangement, substrate binding, and enzyme action. We found that all reported mutations are associated with the active sites or secondary structure interfaces of GNE/MNK. The Persian-Jewish HIBM founder mutation p.M712T is located at the interface alpha4alpha10 and likely affects GlcNAc, Mg2+, and ATP binding. This work contributes to further understanding of GNE/MNK function and ligand binding, which may assist future studies for therapeutic options that target misfolded GNE/MNK in HIBM and/or sialuria.
Topics: Amino Acid Sequence; Humans; Models, Molecular; Molecular Sequence Data; Multienzyme Complexes; Muscular Diseases; Mutation; N-Acetylneuraminic Acid; Protein Conformation; Sialic Acid Storage Disease
PubMed: 19917666
DOI: 10.1093/glycob/cwp176 -
Molecular Genetics and Metabolism Jun 2016Sialuria, a rare inborn error of metabolism, was diagnosed in a healthy 12-year-old boy through whole exome sequencing. The patient had experienced mild delays of speech...
UNLABELLED
Sialuria, a rare inborn error of metabolism, was diagnosed in a healthy 12-year-old boy through whole exome sequencing. The patient had experienced mild delays of speech and motor development, as well as persistent hepatomegaly. Identification of the 8th individual with this disorder, prompted follow-up of the mother-son pair of patients diagnosed over 15years ago. Hepatomegaly was confirmed in the now 19-year-old son, but in the 46-year-old mother a clinically silent liver tumor was detected by ultrasound and MRI. The tumor was characterized as an intrahepatic cholangiocarcinoma (IHCC) and DNA analysis of both tumor and normal liver tissue confirmed the original GNE mutation. As the maternal grandmother in the latter family died at age 49years of a liver tumor, a retrospective study of the remaining pathology slides was conducted and confirmed it to have been an IHCC as well. The overall observation generated the hypothesis that sialuria may predispose to development of this form of liver cancer. As proof of sialuria in the grandmother could not be obtained, an alternate cause of IHCC cannot be ruled out. In a series of 102 patients with IHCC, not a single instance was found with the allosteric site mutation in the GNE gene. This confirms that sialuria is rare even in a selected group of patients, but does not invalidate the concern that sialuria may be a risk factor for IHCC.
SYNOPSIS
Sialuria is a rare inborn error of metabolism characterized by excessive synthesis and urinary excretion of free sialic acid with only minimal clinical morbidity in early childhood, but may be a risk factor for intrahepatic cholangiocarcinoma in adulthood.
Topics: Bile Duct Neoplasms; Child; Cholangiocarcinoma; Female; Hepatomegaly; Heterozygote; Humans; Liver; Liver Neoplasms; Male; Middle Aged; N-Acetylneuraminic Acid; Rare Diseases; Retrospective Studies; Risk Factors; Sialic Acid Storage Disease; Exome Sequencing; Young Adult
PubMed: 27142465
DOI: 10.1016/j.ymgme.2016.04.004 -
The Journal of Biological Chemistry Apr 1991Sialuria is a rare inborn error of metabolism caused by excessive synthesis of sialic acid (N-acetylneuraminic acid, NeuAc). Fibroblasts cultured from the three known...
Sialuria is a rare inborn error of metabolism caused by excessive synthesis of sialic acid (N-acetylneuraminic acid, NeuAc). Fibroblasts cultured from the three known cases of sialuria contained 70-200-fold increases in soluble sialic acid, but normal concentrations of bound sialic acid. The sialic acid appeared in the cytosolic fraction of the cells on differential centrifugation, and was susceptible to borohydride reduction, suggesting that accumulated sialic acid was in the form of NeuAc and not CMP-NeuAc. In biochemical studies, CMP-NeuAc (50 microM) inhibited the UDP-N-acetylglucosamine (UDP-GlcNAc) 2-epimerase of normal fibroblasts by 84-100%, but inhibited the epimerase from sialuria cells by only 19-31%. Feeding sialuria cells up to 5 mM D-glucosamine for 72 h increased free sialic acid content 20-60%, but normal cells were unaffected by this treatment. Cytidine feeding (5 mM, 72 h) reduced the NeuAc content of sialuria cells, initially 112, 104, and 266 nmol/mg protein, by 63-71 nmol/mg protein; CMP-NeuAc concentrations, initially 4, 2, and 5 nmol/mg protein, increased by 14-33 nmol/mg protein. Consequently, the total cellular content of soluble sialic acid (NeuAc + CMP-NeuAc) was lowered 14-46% by cytidine feeding. The inheritance pattern of sialuria has not been determined. However, cells from both parents of one sialuria patient contained normal concentrations of free sialic acid, and the parental epimerase activity also responded normally to CMP-NeuAc. We conclude that the basic biochemical defect in all known cases of sialuria is a failure of CMP-NeuAc to feedback-inhibit UDP-GlcNAc 2-epimerase and cytidine feeding can lower the intracellular soluble sialic acid concentration of sialuria cells.
Topics: Child, Preschool; Chromatography, High Pressure Liquid; Cytidine; Female; Fibroblasts; Humans; Male; Metabolism, Inborn Errors; N-Acetylneuraminic Acid; Sialic Acids
PubMed: 2019577
DOI: No ID Found -
Thrombosis and Haemostasis Jul 2022The gene encodes an enzyme that initiates and regulates the biosynthesis of -acetylneuraminic acid, a precursor of sialic acids. GNE mutations are classically...
The gene encodes an enzyme that initiates and regulates the biosynthesis of -acetylneuraminic acid, a precursor of sialic acids. GNE mutations are classically associated with Nonaka myopathy and sialuria, following an autosomal recessive and autosomal dominant inheritance pattern. Reports show that single GNE variants cause severe thrombocytopenia without muscle weakness. Using panel sequencing, we identified two compound heterozygous variants in in a young girl with life-threatening bleedings, severe congenital thrombocytopenia, and a platelet secretion defect. Both variants are located in the nucleotide-binding site of the -acetylmannosamin kinase domain of GNE. Lectin array showed decreased α-2,3-sialylation on platelets, consistent with loss of sialic acid synthesis and indicative of rapid platelet clearance. Hematopoietic stem cell transplantation (HSCT) normalized platelet counts. This is the first report of an HSCT in a patient with an inherited GNE defect leading to normal platelet counts.
Topics: Blood Platelets; Distal Myopathies; Female; Humans; Multienzyme Complexes; Mutation; N-Acetylneuraminic Acid; Thrombocytopenia
PubMed: 35052006
DOI: 10.1055/s-0041-1742207 -
Pediatric Neurology Nov 2023Biallelic pathogenic variants in SLC17A5 cause three forms of free sialic acid storage disease categorized based on severity from least to most severe: Salla disease,...
BACKGROUND
Biallelic pathogenic variants in SLC17A5 cause three forms of free sialic acid storage disease categorized based on severity from least to most severe: Salla disease, intermediate-severe Salla disease, and infantile free sialic acid storage disease. Intermediate-severe Salla disease is the most recently described form. Here, we report a longitudinal characterization of intermediate-severe Salla disease progression in two sisters carrying the following biallelic variants in SLC17A5: c.406A>G (p.Lys136Glu) and c.819+1G>A.
METHODS
A retrospective review of medical records was performed. A developmental questionnaire was completed to obtain further clinical information. For functional characterization of the predicted splice site variant, RNA was extracted from patient blood samples and sequenced.
RESULTS
Disease onset occurred within the first six months of life in both patients. Early childhood development was delayed with achievement of some milestones followed by a developmental plateau in late childhood. After this, both patients began a slow and progressive neurological regression in adolescence. Functional studies confirmed the pathogenicity of the c.819+1G>A variant, resulting in a frameshift and deletion of exon 6.
CONCLUSIONS
We present a detailed study describing the clinical course of intermediate-severe Salla disease with over 15 to 20 years of evolution and demonstrate the pathogenicity of the c.819+1G>A splice site variant.
Topics: Adolescent; Humans; Child; Child, Preschool; Sialic Acid Storage Disease; Mutation; N-Acetylneuraminic Acid; Disease Progression
PubMed: 37713976
DOI: 10.1016/j.pediatrneurol.2023.08.013 -
Biochimica Et Biophysica Acta Jun 2006Sialuria is an inborn error of metabolism characterized by coarse face, hepatomegaly and recurrent respiratory tract infections. The genetic defect in this disorder...
Sialuria is an inborn error of metabolism characterized by coarse face, hepatomegaly and recurrent respiratory tract infections. The genetic defect in this disorder results in a loss of feedback control of UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine-kinase by CMP-N-acetylneuraminic acid (CMP-NeuAc) resulting in a substantial overproduction of cytoplasmic free sialic acid. This study addresses fibroblast CMP-NeuAc levels and N- and O-glycan sialylation of serum proteins from Sialuria patients. CMP-NeuAc levels were measured with HPLC in fibroblasts. Isoelectric focusing (IEF) of serum transferrin and of apolipoprotein C-III (apoC-III) was performed on serum of three Sialuria patients. Isoforms of these proteins can be used as specific markers for the biosynthesis of N- and core 1 O-glycans. Furthermore, total N- and O-linked glycans from serum proteins were analyzed by HPLC. HPLC showed a clear overproduction of CMP-NeuAc in fibroblasts of a Sialuria patient. Minor changes were found for serum N-glycans and hypersialylation was found for core 1 O-glycans on serum apoC-III and on total serum O-glycans in Sialuria patients. HPLC showed an increased ratio of disialylated over monosialylated core 1 O-glycans. The hypersialylation of core 1 O-glycans is due to the increase of NeuAcalpha2,6-containing structures (mainly NeuAcalpha2-3Galbeta1-3[NeuAcalpha2-6]GalNAc). This may relate to KM differences between GalNAc-alpha2,6-sialyltransferase and alpha2,3-sialyltransferases. This is the first study demonstrating that the genetic defect in Sialuria results in a CMP-NeuAc overproduction. Subsequently, increased amounts of alpha2,6-linked NeuAc were found on serum core 1 O-glycans from Sialuria patients. N-glycosylation of serum proteins seems largely unaffected. Sialuria is the first metabolic disorder presenting with hypersialylated O-glycans.
Topics: Apolipoprotein C-III; Apolipoproteins C; Chromatography, High Pressure Liquid; Glycoproteins; Glycosylation; Humans; Isoelectric Focusing; N-Acetylneuraminic Acid; Nucleotides; Polysaccharides; Protein Isoforms; Sialic Acid Storage Disease; Transferrin
PubMed: 16769205
DOI: 10.1016/j.bbadis.2006.03.009 -
American Journal of Human Genetics Jun 2001"French type" sialuria, a presumably dominant disorder that, until now, had been documented in only five patients, manifests with mildly coarse facies, slight motor...
"French type" sialuria, a presumably dominant disorder that, until now, had been documented in only five patients, manifests with mildly coarse facies, slight motor delay, and urinary excretion of large quantities (>1 g/d) of free N-acetylneuraminic acid (NeuAc). The basic defect consists of the very rare occurrence of failed feedback inhibition of a rate-limiting enzyme, in this case uridinediphosphate-N-acetylglucosamine (UDP-GlcNAc) 2-epimerase, by a downstream product, in this case cytidine monophosphate (CMP)-NeuAc. We report a new patient with sialuria who has a heterozygous G-->A substitution in nucleotide 848 of the epimerase gene, which results in an R266Q change. The proband's other allele, as expected, had no mutation. However, the heterozygous R266Q mutation was detected in the patient's mother, who has similarly increased urinary levels of free NeuAc, thereby confirming, for the first time, the dominant mode of inheritance of this inborn error. The biochemical diagnosis of the proband was verified by the greatly increased level of free NeuAc in his cultured fibroblasts, the NeuAc distribution, mainly (59%) in the cytoplasm, and by the complete failure of 100 microM CMP-NeuAc to inhibit UDP-GlcNAc 2-epimerase activity in the mutant cells. These findings call for expansion of the phenotype to include adults and for more-extensive assaying of free NeuAc in the urine of children with mild developmental delay. The prevalence of sialuria is probably grossly underestimated.
Topics: Adult; Base Sequence; Carbohydrate Epimerases; Child; Child, Preschool; Cytidine Monophosphate N-Acetylneuraminic Acid; Cytoplasm; Developmental Disabilities; Escherichia coli Proteins; Feedback; Female; Fibroblasts; France; Genes, Dominant; Heterozygote; Humans; Infant; Infant, Newborn; Male; Metabolism, Inborn Errors; Middle Aged; Molecular Sequence Data; Mutation, Missense; Pedigree; Sialic Acids; Uridine Diphosphate N-Acetylglucosamine
PubMed: 11326336
DOI: 10.1086/320598 -
Annals of Neurology Jun 2009We performed high-resolution in vitro proton nuclear magnetic resonance spectroscopy on cerebrospinal fluid and urine samples of 44 patients with leukodystrophies of... (Comparative Study)
Comparative Study
We performed high-resolution in vitro proton nuclear magnetic resonance spectroscopy on cerebrospinal fluid and urine samples of 44 patients with leukodystrophies of unknown cause. Free sialic acid concentration was increased in cerebrospinal fluid of two siblings with mental retardation and mild hypomyelination. By contrast, urinary excretion of free sialic acid in urine was normal on repeated testing by two independent methods. Both patients were homozygous for the K136E mutation in SLC17A5, the gene responsible for the free sialic acid storage diseases. Our findings demonstrate that mutations in the SLC17A5 gene have to be considered in patients with hypomyelination, even in the absence of sialuria.
Topics: Adolescent; Child; Diagnosis, Differential; Hereditary Central Nervous System Demyelinating Diseases; Humans; N-Acetylneuraminic Acid; Nuclear Magnetic Resonance, Biomolecular; Organic Anion Transporters; Sialic Acid Storage Disease; Symporters; Young Adult
PubMed: 19557856
DOI: 10.1002/ana.21624 -
Experimental Neurology May 2017Slc17a5 mice represent an animal model for the infantile form of sialic acid storage disease (SASD). We analyzed genetic and histological time-course expression of...
Slc17a5 mice represent an animal model for the infantile form of sialic acid storage disease (SASD). We analyzed genetic and histological time-course expression of myelin and oligodendrocyte (OL) lineage markers in different parts of the CNS, and related this to postnatal neurobehavioral development in these mice. Sialin-deficient mice display a distinct spatiotemporal pattern of sialic acid storage, CNS hypomyelination and leukoencephalopathy. Whereas few genes are differentially expressed in the perinatal stage (p0), microarray analysis revealed increased differential gene expression in later postnatal stages (p10-p18). This included progressive upregulation of neuroinflammatory genes, as well as continuous down-regulation of genes that encode myelin constituents and typical OL lineage markers. Age-related histopathological analysis indicates that initial myelination occurs normally in hindbrain regions, but progression to more frontal areas is affected in Slc17a5 mice. This course of progressive leukoencephalopathy and CNS hypomyelination delays neurobehavioral development in sialin-deficient mice. Slc17a5 mice successfully achieve early neurobehavioral milestones, but exhibit progressive delay of later-stage sensory and motor milestones. The present findings may contribute to further understanding of the processes of CNS myelination as well as help to develop therapeutic strategies for SASD and other myelination disorders.
Topics: Age Factors; Animals; Animals, Newborn; Brain; Developmental Disabilities; Disease Models, Animal; Gene Expression Regulation, Developmental; Glial Fibrillary Acidic Protein; Intermediate Filaments; Leukoencephalopathies; Lysosomal-Associated Membrane Protein 1; Mental Disorders; Mice; Mice, Inbred C57BL; Mice, Transgenic; Organic Anion Transporters; Sialic Acid Storage Disease; Symporters
PubMed: 28189729
DOI: 10.1016/j.expneurol.2017.02.009 -
Scientific Reports May 2018Sialic acids (SAs) are nine carbon acidic amino sugars, found at the outermost termini of glycoconjugates performing various physiological and pathological functions. SA...
Sialic acids (SAs) are nine carbon acidic amino sugars, found at the outermost termini of glycoconjugates performing various physiological and pathological functions. SA synthesis is regulated by UDP-GlcNAc 2-epimerase/ManNAc kinase (GNE) that catalyzes rate limiting steps. Mutations in GNE result in rare genetic disorders, GNE myopathy and Sialuria. Recent studies indicate an alternate role of GNE in cell apoptosis and adhesion, besides SA biosynthesis. In the present study, using a HEK cell-based model for GNE myopathy, the role of Insulin-like Growth Factor Receptor (IGF-1R) as cell survival receptor protein was studied to counter the apoptotic effect of non-functional GNE. In the absence of functional GNE, IGF-1R was hyposialylated and transduced a downstream signal upon IGF-1 (IGF-1R ligand) treatment. IGF-1 induced activation of IGF-1R led to AKT (Protein Kinase B) phosphorylation that may phosphorylate BAD (BCL2 Associated Death Promoter) and its dissociation from BCL2 to prevent apoptosis. However, reduced ERK (Extracellular signal-regulated kinases) phosphorylation in GNE deficient cells after IGF-1 treatment suggests downregulation of the ERK pathway. A balance between the ERK and AKT pathways may determine the cell fate towards survival or apoptosis. Our study suggests that IGF-1R activation may rescue apoptotic cell death of GNE deficient cell lines and has potential as therapeutic target.
Topics: Apoptosis; Carbohydrate Epimerases; Gene Knockdown Techniques; HEK293 Cells; Humans; Mitochondria; N-Acetylneuraminic Acid; Protein Transport; Receptor, IGF Type 1; Receptors, Somatomedin
PubMed: 29743626
DOI: 10.1038/s41598-018-25510-9