A Gene on Chromosome 11q23 Coding for a Putative Glucose- 6-Phosphate Translocase Is Mutated in Glycogen-Storage Disease Types Ib and Ic (original) (raw)

The putative glucose 6-phosphate translocase gene is mutated in essentially all cases of glycogen storage disease type I non-a

European Journal of Human Genetics, 1999

The purpose of this work was to test the hypothesis that mutations in the putative glucose 6-phosphate translocase gene would account for most of the cases of GSD I that are not explained by mutations in the phosphohydrolase gene, ie that are not type Ia. Twenty-three additional families diagnosed as having GSD I non-a (GSD Ib, Ic or Id) have now been analysed. The 9 exons of the gene were amplified by PCR and mutations searched both by SSCP and heteroduplex analysis. Except for one family in which only one mutation was found, all patients had two allelic mutations in the gene encoding the putative glucose 6-phosphate translocase. Sixteen of the mutations are new and they are all predicted to lead to nonfunctional proteins. All investigated patients had some degree of neutropenia or neutrophil dysfunction and the clinical phenotype of the four new patients who had been diagnosed as GSD Ic and the one diagnosed as GSD Id was no different from the GSD Ib patients. Since these patients, and the four type Ic patients from two families previously studied, shared several mutations with GSD Ib patients, we conclude that their basic defect is in the putative glucose 6-phosphate translocase and that they should be reclassified as GSD Ib. Isolated defects in microsomal Pi transporter or in microsomal glucose transporter must be very rare or have phenotypes that are not recognised as GSD I, so that in practice there are only two subtypes of GSD I (GSD Ia and GSD Ib).

Type-1c glycogen storage disease is not caused by mutations in the glucose-6-phosphate transporter gene

Human Genetics, 1999

Glycogen storage disease type 1 (GSD-1) is a group of autosomal recessive disorders caused by deficiencies in glucose-6-phosphatase (G6Pase) and the associated substrate/product transporters. Molecular genetic studies have demonstrated that GSD-1a and GSD-1b are caused by mutations in the G6Pase enzyme and a glucose-6-phosphate transporter (G6PT), respectively. While kinetic studies of G6Pase catalysis predict that the index GSD-1c patient is deficient in a pyrophosphate/phosphate transporter, the existence of a separate locus for GSD-1c remains unclear. We have previously shown that the G6Pase gene of the index GSD-1c patient is intact; we now show that the G6PT gene of this patient is normal, strongly suggesting the existence of a distinct GSD-1c locus.

Mutations in the glucose-6-phosphate transporter (G6PT) gene in patients with glycogen storage diseases type 1b and 1c

FEBS Letters, 1999

Glycogen storage diseases type 1 (GSD 1) are a group of autosomal recessive disorders characterized by impairment of terminal steps of glycogenolysis and gluconeogenesis. Mutations of the glucose-6-phosphatase gene are responsible for the most frequent form of GSD 1, the subtype 1a, while mutations of the glucose-6-phosphate transporter gene (G6PT) have recently been shown to cause the non 1a forms of GSD, namely the 1b and 1c subtypes. Here, we report on the analysis by single-stranded conformation polymorphism (SSCP) and/or DNA sequencing of the exons of the G6PT in 14 patients diagnosed either as affected by the GSD 1b or 1c subtypes. Mutations in the G6PT gene were found in all patients. Four of the detected mutations were novel mutations, while the others were previously described. Our results confirm that the GSD 1b and 1c forms are due to mutations in the same gene, i.e. the G6PT gene. We also show that the same kind of mutation can be associated or not with evident clinical complications such as neutrophil impairment. Since no correlation between the type and position of the mutation and the severity of the disease was found, other unknown factors may cause the expression of symptoms, such as neutropenia, which dramatically influence the severity of the disease.

The glucose-6-phosphate transporter is a phosphate-linked antiporter deficient in glycogen storage disease type Ib and Ic

The FASEB Journal, 2008

Glycogen storage disease type Ib (GSD-Ib) is caused by deficiencies in the glucose-6-phosphate (G6P) transporter (G6PT) that have been well characterized. Interestingly, deleterious mutations in the G6PT gene were identified in clinical cases of GSD type Ic (GSD-Ic) proposed to be deficient in an inorganic phosphate (P i ) transporter. We hypothesized that G6PT is both the G6P and P i transporter. Using reconstituted proteoliposomes we show that both G6P and P i are efficiently taken up into P i -loaded G6PTproteoliposomes. The G6P uptake activity decreases as the internal:external P i ratio decreases and the P i uptake activity decreases in the presence of external G6P. Moreover, G6P or P i uptake activity is not detectable in P i -loaded proteoliposomes containing the p.R28H G6PT null mutant. The G6PT-proteoliposomemediated G6P or P i uptake is inhibited by cholorgenic acid and vanadate, both specific G6PT inhibitors. Glucose-6-phosphatase-␣ (G6Pase-␣), which facilitates microsomal G6P uptake by G6PT, fails to stimulate G6P uptake in P i -loaded G6PT-proteoliposomes, suggesting that the G6Pase-␣-mediated stimulation is caused by decreasing G6P and increasing P i concentrations in microsomes. Taken together, our results suggest that G6PT has a dual role as a G6P and a P i transporter and that GSD-Ib and GSD-Ic are deficient in the same G6PT gene.-Chen, S.-Y., Pan, C.-.J, Nandigama, K., Mansfield, B., Ambudkar, S., Chou, J. The glucose-6-phosphate transporter is a phosphate-linked antiporter deficient in glycogen storage disease type Ib and Ic. FASEB J. 22, 2206 -2213 (2008)

NPT4, a new microsomal phosphate transporter: Mutation analysis in glycogen storage disease type Ic

Journal of Inherited Metabolic Disease, 2004

De¢ciency of a microsomal phosphate transporter in the liver has been suggested in some patients affected by glycogen storage disease type Ic (GSD Ic). Several Na þ /phosphate co-transporters have been characterized as members of the anion^cation symporter family. Recently, the cDNA sequence of two phosphate transporters, NPT3 and NPT4, expressed in liver, kidney and intestine, has been determined. We studied expression of human NPT4 in COS cells and observed an ER localization of the transporter by immuno-£uorescence microscopy. We speculated that this transporter could play a role in the regulation of the glucose-6-phosphatase (G6-Pase) complex. We revealed the genomic structure of NPT4 and analysed the gene as a candidate for GSD Ic. DNA was collected from ¢ve patients without mutations in G6-Pase or the G6-P transporter gene. DNA analysis of NPT4 revealed that one patient was heterozygous for a G>A transition at nucleotide 601 which would result in a G201R substitution. Our results do not con¢rm the hypothesis that this gene is mutated in GSD Ic patients. However, we cannot exclude that the mutation found reduces the phosphate transport ef¢ciency, possibly modulating the G6-Pase complex. Glycogen storage disease type I (GSD I; McKusick 232200) is an autosomal recessive disorder with an incidence of 1 in 100 000 live births. GSD I is caused by the de¢ciency of the glucose-6-phosphatase system (G6-Pase) and the associated transporters in liver, kidney and intestine. G6-Pase catalyses the hydrolysis of glucose 6-phosphate to glucose and phosphate and the active site is situated inside

Structure-function analysis of the glucose-6-phosphate transporter deficient in glycogen storage disease type Ib

Human Molecular Genetics, 2002

Glycogen storage disease type Ib (GSD-Ib) is caused by a deficiency in the glucose-6-phosphate transporter (G6PT), a 10 transmembrane domain endoplasmic reticulum protein. To date, 69 G6PT mutations, including 28 missenses and 2 codon deletions, have been identified in GSD-Ib patients. We previously characterized 15 of the missense and one codon deletion mutations using a pSVL-based expression assay. A lack of sensitivity in this assay limited the discrimination between mutations that lead to loss of function and mutations that leave a low residual activity. We now report an improved G6PT assay, based on an adenoviral vector-mediated expression system and its use in the functional characterization of all 30 codon mutations found in GSD-Ib patients. Twenty of the naturally occurring mutations completely abolish microsomal G6P uptake activity while the other 10 mutations, including 5 previously characterized ones, partially inactivate the transporter. This information should greatly facilitate genotype-phenotype correlation. We also report a structurefunction analysis of G6PT. In addition to the 3 destabilizing mutations reported previously, we now show that the G50R, C176R, V235del, G339C and G339D mutations also compromise the G6PT stability. Mutation analysis of the amino-terminal domain of G6PT shows that it is required for optimal G6P uptake activity. Finally, we show that degradation of both wild-type and mutant G6PT is inhibited by a potent proteasome inhibitor, lactacystin, demonstrating that G6PT is a substrate for proteasome-mediated degradation.

Identification, purification and genetic deficiencies of the glucose-6-phosphatase system transport proteins

European Journal of Pediatrics, 1993

Hepatic microsomal glucose-6-phosphatase (Glc-6-P'ase) is a complex multicomponent system containing at least three transport proteins, in addition to the catalytic subunit and a Ca 2+ binding regulatory protein. The transport proteins have been designated Tt the glucose-6-phosphate transport protein, T2 a phosphate/ pyrophosphate transport protein and T3 a glucose transport protein. Diagnosis of the genetic deficiencies of these transport proteins at present requires a complex kinetic analysis of the Glc-6-P'ase system as a whole.

Homology Modeling of the Human Microsomal Glucose 6-Phosphate Transporter Explains the Mutations That Cause the Glycogen Storage Disease Type Ib †

Biochemistry, 2004

Glycogen storage disease type Ib is caused by mutations in the glucose 6-phosphate transporter (G6PT) in the endoplasmic reticulum membrane in liver and kidney. Twenty-eight missense and two deletion mutations that cause the disease were previously shown to reduce or abolish the transporter's activity. However, the mechanisms by which these mutations impair transport remain unknown. On the basis of the recently determined crystal structure of its Escherichia coli homologue, the glycerol 3-phosphate transporter, we built a three-dimensional structural model of human G6PT by homology modeling. G6PT is proposed to consist of 12 transmembrane R-helices that are divided into N-and C-terminal domains, with the substrate-translocation pore located between the two domains and the substrate-binding site formed by R28 and K240 at the domain interface. The disease-causing mutations were found to occur at four types of positions: (I) in the substrate-translocation pore, (II) at the N-/C-terminal domain interface, (III) in the interior of the N-and C-terminal domains, and (IV) on the protein surface. Whereas class I mutations affect substrate binding directly, class II mutations, mostly involving changes in side chain size, charge, or both, hinder the conformational change required for substrate translocation. On the other hand, class III and class IV mutations, often introducing a charged residue into a helix bundle or at the protein-lipid interface, probably destabilize the protein. These results also suggest that G6PT operates by a similar antiport mechanism as its E. coli homologue, namely, the substrate binds at the N-and C-terminal domain interface and is then transported across the membrane via a rocker-switch type of movement of the two domains.