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Janice Y. Chou - One of the best experts on this subject based on the ideXlab platform.
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type i glycogen storage diseases disorders of the glucose 6 phosphatase glucose 6 Phosphate Transporter complexes
Journal of Inherited Metabolic Disease, 2015Co-Authors: Janice Y. Chou, Brian C. Mansfield, Hyun Sik JunAbstract:Disorders of the glucose-6-phosphatase (G6Pase)/glucose-6-Phosphate Transporter (G6PT) complexes consist of three subtypes: glycogen storage disease type Ia (GSD-Ia), deficient in the liver/kidney/intestine-restricted G6Pase-α (or G6PC); GSD-Ib, deficient in a ubiquitously expressed G6PT (or SLC37A4); and G6Pase-β deficiency or severe congenital neutropenia syndrome type 4 (SCN4), deficient in the ubiquitously expressed G6Pase-β (or G6PC3). G6Pase-α and G6Pase-β are glucose-6-Phosphate (G6P) hydrolases with active sites lying inside the endoplasmic reticulum (ER) lumen and as such are dependent upon the G6PT to translocate G6P from the cytoplasm into the lumen. The tissue expression profiles of the G6Pase enzymes dictate the disease's phenotype. A functional G6Pase-α/G6PT complex maintains interprandial glucose homeostasis, while a functional G6Pase-β/G6PT complex maintains neutrophil/macrophage energy homeostasis and functionality. G6Pase-β deficiency is not a glycogen storage disease but biochemically it is a GSD-I related syndrome (GSD-Irs). GSD-Ia and GSD-Ib patients manifest a common metabolic phenotype of impaired blood glucose homeostasis not shared by GSD-Irs. GSD-Ib and GSD-Irs patients manifest a common myeloid phenotype of neutropenia and neutrophil/macrophage dysfunction not shared by GSD-Ia. While a disruption of the activity of the G6Pase-α/G6PT complex readily explains why GSD-Ia and GSD-Ib patients exhibit impaired glucose homeostasis, the basis for neutropenia and myeloid dysfunction in GSD-Ib and GSD-Irs are only now starting to be understood. Animal models of all three disorders are now available and are being exploited to both delineate the disease more precisely and develop new treatment approaches, including gene therapy.
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functional analysis of mutations in the glucose 6 Phosphate Transporter that cause glycogen storage disease type ib
Molecular Genetics and Metabolism, 2008Co-Authors: Shihyin Chen, Chijiunn Pan, Soojung Lee, Wentao Peng, Janice Y. ChouAbstract:Abstract The glucose-6-Phosphate Transporter (G6PT) deficient in glycogen storage disease type Ib is a Phosphate (P i )-linked antiporter capable of G6P: P i and P i :P i exchanges. We previously characterized G6PT mutations by measuring G6P uptake activities in microsomes co-expressing G6PT and glucose-6-phosphatase-α. Here we report a new assay, based on reconstituted proteoliposomes carrying only G6PT, and characterize G6P and P i uptake activities of 23 G6PT mutations. We show that co-expression and G6PT-only assays are equivalent in measuring G6PT activity. However, the p.Q133P mutation exhibits differential G6P and P i transport activities, suggesting that characterizing G6P and P i transport activities of G6PT mutations may yield insights to this genetic disorder.
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structure function analysis of the glucose 6 Phosphate Transporter deficient in glycogen storage disease type ib
Human Molecular Genetics, 2002Co-Authors: Liyuan Chen, Chijiunn Pan, Jengjer Shieh, Janice Y. ChouAbstract: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 structure– function 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.
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inactivation of the glucose 6 Phosphate Transporter causes glycogen storage disease type 1b
Journal of Biological Chemistry, 1999Co-Authors: Hisayuki Hiraiwa, Baochuan Lin, Chijiunn Pan, S W Moses, Janice Y. ChouAbstract:Glycogen storage disease type 1b (GSD-1b) is proposed to be caused by a deficiency in microsomal glucose 6-Phosphate (G6P) transport, causing a loss of glucose-6-phosphatase activity and glucose homeostasis. However, for decades, this disorder has defied molecular characterization. In this study, we characterize the structural organization of the G6P Transporter gene and identify mutations in the gene that segregate with the GSD-1b disorder. We report the functional characterization of the recombinant G6P Transporter and demonstrate that mutations uncovered in GSD-1b patients disrupt G6P transport. Our results, for the first time, define a molecular basis for functional deficiency in GSD-1b and raise the possibility that the defective G6P Transporter contributes to neutropenia and neutrophil/monocyte dysfunctions characteristic of GSD-1b patients.
Yoshinori Moriyama - One of the best experts on this subject based on the ideXlab platform.
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type 1 sodium dependent Phosphate Transporter acts as a membrane potential driven urate exporter
Current Molecular Pharmacology, 2013Co-Authors: Takaaki Miyaji, Tatsuya Kawasaki, Natsuko Togawa, Hiroshi Omote, Yoshinori MoriyamaAbstract:SLC17A1 protein (NPT1) was the first identified member of the SLC17 Phosphate Transporter family, and is known to mediate Na + /inorganic Phosphate (Pi) co-transport when expressed in Xenopus oocytes. Although this protein was suggested to be a renal polyspecific anion exporter, its transport properties were not well characterized. The clean biochemical approach revealed that proteoliposomes comprising purified NPT1 as the only protein source transport various organic anions such as urate, p-aminohippuric acid (PAH), and acetylsalicylic acid (aspirin) in a membrane potential (Δψ)-driven and Cl - -dependent manner. Human NPT1 carrying an SNP mutation, Thr269Ile, known to increase the risk of gout, exhibited 32% lower urate transport activity compared to the wild type protein, leading to the conclusion that NPT1 is the long searched for Transporter responsible for renal urate excretion. In the present article, we summarized the history of identification of the urate exporter and its possible involvement in the dynamism of urate under physiological and pathological conditions.
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type 1 sodium dependent Phosphate Transporter slc17a1 protein is a cl dependent urate exporter
Journal of Biological Chemistry, 2010Co-Authors: Masafumi Iharada, Takaaki Miyaji, Hiroshi Omote, Takahiro Fujimoto, Miki Hiasa, Naohiko Anzai, Yoshinori MoriyamaAbstract:Abstract SLC17A1 protein (NPT1) is the first identified member of the SLC17 Phosphate Transporter family and mediates the transmembrane cotransport of Na+/Pi in oocytes. Although this protein is believed to be a renal polyspecific anion exporter, its transport properties are not well characterized. Here, we show that proteoliposomes containing purified SLC17A1 transport various organic anions such as p-aminohippuric acid and acetylsalicylic acid (aspirin) in an inside positive membrane potential (Δψ)-dependent manner. We found that NPT1 also transported urate. The uptake characteristics were similar to that of SLC17 members in its Cl− dependence and inhibitor sensitivity. When arginine 138, an essential amino acid residue for members of the SLC17 family such as the vesicular glutamate Transporter, was specifically mutated to alanine, the resulting mutant protein was inactive in Δψ-dependent anion transport. Heterologously expressed and purified human NPT1 carrying the single nucleotide polymorphism mutation that is associated with increased risk of gout in humans exhibited 32% lower urate transport activity compared with the wild type protein. These results strongly suggested that NPT1 is a Cl−-dependent polyspecific anion exporter involved in urate excretion under physiological conditions.
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type 1 sodium dependent Phosphate Transporter slc17a1 protein is a cl dependent urate exporter
Journal of Biological Chemistry, 2010Co-Authors: Masafumi Iharada, Takaaki Miyaji, Hiroshi Omote, Takahiro Fujimoto, Miki Hiasa, Naohiko Anzai, Yoshinori MoriyamaAbstract:SLC17A1 protein (NPT1) is the first identified member of the SLC17 Phosphate Transporter family and mediates the transmembrane cotransport of Na(+)/P(i) in oocytes. Although this protein is believed to be a renal polyspecific anion exporter, its transport properties are not well characterized. Here, we show that proteoliposomes containing purified SLC17A1 transport various organic anions such as p-aminohippuric acid and acetylsalicylic acid (aspirin) in an inside positive membrane potential (Deltapsi)-dependent manner. We found that NPT1 also transported urate. The uptake characteristics were similar to that of SLC17 members in its Cl(-) dependence and inhibitor sensitivity. When arginine 138, an essential amino acid residue for members of the SLC17 family such as the vesicular glutamate Transporter, was specifically mutated to alanine, the resulting mutant protein was inactive in Deltapsi-dependent anion transport. Heterologously expressed and purified human NPT1 carrying the single nucleotide polymorphism mutation that is associated with increased risk of gout in humans exhibited 32% lower urate transport activity compared with the wild type protein. These results strongly suggested that NPT1 is a Cl(-)-dependent polyspecific anion exporter involved in urate excretion under physiological conditions.
Hiroko Segawa - One of the best experts on this subject based on the ideXlab platform.
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evidence of an intestinal Phosphate Transporter alternative to type iib sodium dependent Phosphate Transporter in rats with chronic kidney disease
Nephrology Dialysis Transplantation, 2021Co-Authors: Yasuhiro Ichida, Hiroko Segawa, Kenichi Miyamoto, Shuichi Ohtomo, Tessai Yamamoto, Naoaki Murao, Yoshinori Tsuboi, Yoshiki Kawabe, Naoshi Horiba, Jurgen FloegeAbstract:Background Phosphate is absorbed in the small intestine via passive flow and active transport.NaPi-IIb, a type II sodium-dependent Phosphate Transporter, is considered to mediate active Phosphate transport in rodents. To study the regulation of intestinal Phosphate transport in chronic kidney disease (CKD), we analyzed the expression levels of NaPi-IIb, pituitary-specific transcription factor 1 (PiT-1) and PiT-2 and the kinetics of intestinal Phosphate transport using two CKD models. Methods CKD was induced in rats via adenine orThy1 antibody injection. Phosphate uptake by intestinal brush border membrane vesicles (BBMV) and the messenger RNA (mRNA) expression of NaPi-IIb, PiT-1 and PiT-2 were analyzed. The protein expression level of NaPi-IIb was measured by mass spectrometry (e.g. liquid chromatography tandem mass spectrometry). Results In normal rats, Phosphate uptake into BBMV consisted of a single saturable component and its Michaelis constant (Km) was comparable to that of NaPi-IIb. The maximum velocity (Vmax) correlated with mRNA and protein levels of NaPi-IIb. In the CKD models, intestinal Phosphate uptake consisted of two saturable components. The Vmax of the higher-affinity transport, which is thought to be responsible for NaPi-IIb, significantly decreased and the decrease correlated with reduced NaPi-IIb expression. The Km of the lower-affinity transport was comparable to that of PiT-1 and -2. PiT-1 mRNA expression was much higher than that of PiT-2, suggesting that PiT-1 was mostly responsible for Phosphate transport. Conclusions This study suggests that the contribution of NaPi-IIb to intestinal Phosphate absorption dramatically decreases in rats with CKD and that a low-affinity alternative to NaPi-IIb, in particular PiT-1, is upregulated in a compensatory manner in CKD.
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type iic sodium dependent Phosphate Transporter regulates calcium metabolism
Journal of The American Society of Nephrology, 2009Co-Authors: Hiroko Segawa, Masashi Kuwahata, Fumito Aranami, Akemi Onitsuka, Etsuyo Hanabusa, Junya Furutani, Ichiro Kaneko, Yuka Tomoe, Natsuki Matsumoto, Mitsuru MatsumotoAbstract:Primary renal inorganic Phosphate (Pi) wasting leads to hypoPhosphatemia, which is associated with skeletal mineralization defects. In humans, mutations in the gene encoding the type IIc sodium–dependent Phosphate Transporter lead to hereditary hypophophatemic rickets with hypercalciuria, but whether Pi wasting directly causes the bone disorder is unknown. Here, we generated Npt2c-null mice to define the contribution of Npt2c to Pi homeostasis and to bone abnormalities. Homozygous mutants (Npt2c−/−) exhibited hypercalcemia, hypercalciuria, and elevated plasma 1,25-dihydroxyvitamin D3 levels, but they did not develop hypoPhosphatemia, hyperphosphaturia, renal calcification, rickets, or osteomalacia. The increased levels of 1,25-dihydroxyvitamin D3 in Npt2c−/− mice compared with age-matched Npt2c+/+ mice may be the result of reduced catabolism, because we observed significantly reduced expression of renal 25-hydroxyvitamin D–24-hydroxylase mRNA but no change in 1α-hydroxylase mRNA levels. Enhanced intestinal absorption of calcium (Ca) contributed to the hypercalcemia and increased urinary Ca excretion. Furthermore, plasma levels of the phosphaturic protein fibroblast growth factor 23 were significantly decreased in Npt2c−/− mice. Sodium-dependent Pi co-transport at the renal brush border membrane, however, was not different among Npt2c+/+, Npt2c+/−, and Npt2c−/− mice. In summary, these data suggest that Npt2c maintains normal Ca metabolism, in part by modulating the vitamin D/fibroblast growth factor 23 axis.
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hereditary hypoPhosphatemic rickets with hypercalciuria a study for the Phosphate Transporter gene type iic and osteoblastic function
Journal of Bone and Mineral Metabolism, 2007Co-Authors: Takehisa Yamamoto, Hiroko Segawa, Kenichi Miyamoto, Toshimi Michigami, Fumito Aranami, Shigeo Nakajima, Keiichi OzonoAbstract:Two cases of hereditary hypoPhosphatemic rickets with hypercalciuria (HHRH) were reported in Japanese female siblings. Both of them manifested short stature and bowed legs, and biochemical examination revealed hypoPhosphatemia, phosphaturia, and hypercalciuria. The serum concentrations of 1,25-dihydroxyvitamin D (1,25(OH)2D) were elevated. In the oral Phosphate loading test, serum Phosphate levels were markedly increased in the HHRH patients, and the elevation was much higher than that in patients affected with X-linked hypoPhosphatemic rickets (XLH), suggesting the increased gastrointestinal absorption of Phosphate in HHRH. Bone histology studies showed increased osteoid surface and width in HHRH, which was compatible with osteomalacia. In the HHRH patients, there were no hypomineralized periosteocytic lesions, which was a hallmark of XLH in bone histology. In one of the HHRH patients, Phosphate administration alone almost completely cured the osteomalacia within a year, although pharmacological doses of 1,25(OH)2D3 had little effect. In osteoblasts isolated from a HHRH patient, basal alkaline phosphatase (ALP) activities and osteocalcin syntheses by a physiological concentration of 1,25(OH)2D3 were not stimulated by the increased medium Phosphate concentrations from 0.5 to 4 mM. In contrast, these two parameters were stimulated by the increased medium Phosphate concentrations both in normal and XLH osteoblasts, although the regulatory patterns of increased osteocalcin syntheses were different from normal to XLH osteoblasts; 2 and 4 mM of Phosphate concentrations at least were necessary for normal and XLH osteoblasts, respectively. The gene analysis of Phosphate Transporter revealed a novel heterozygous mutation (R564C) in the exon of Phosphate Transporter NPT type IIc. These lines of evidence suggested that the pathogenesis of osteomalacia in HHRH was different from XLH in terms of the utility of Phosphate in osteoblasts. These abnormalities were speculated to be associated with the abnormal functions of Phosphate Transporter gene type IIc, although the exact roles of this Phosphate Transporter in the human osteoblast are still unknown.
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new aspect of renal Phosphate reabsorption the type iic sodium dependent Phosphate Transporter
American Journal of Nephrology, 2007Co-Authors: Kenichi Miyamoto, Sawako Tatsumi, Mikiko Ito, Masashi Kuwahata, Hiroko SegawaAbstract:Abnormalities of the inorganic Phosphate (Pi) reabsorption in the kidney result in various metabolic disorders. Na+-dependent Pi (Na/Pi) Transporters in the brush border membrane of proximal tubular cells mediate the rate-limiting step in the overall Pi-reabsorptive process. Type IIa and type IIc Na/Pi coTransporters are expressed in the apical membrane of proximal tubular cells and mediate Na/Pi cotransport; the extent of Pi reabsorption in the proximal tubules is determined largely by the abundance of the type IIa Na/Pi coTransporter. However, several studies suggest that the type IIc coTransporter in Pi reabsorption may also play a role in this process. For example, mutation of the type IIc Na/Pi coTransporter gene results in hereditary hypoPhosphatemic rickets with hypercalciuria, suggesting that the type IIc Transporter plays an important role in renal Pi reabsorption in humans and may be a key determinant of the plasma Pi concentration. The type IIc Na/Pi Transporter is regulated by parathyroid hormone, dietary Pi, and fibroblast growth factor 23, and studies suggest a differential regulation of the IIa and IIc Transporters. Indeed, differences in temporal and/or spatial expression of the type IIa and type IIc Na/Pi Transporters may be required for normal Phosphate homeostasis and bone development. This review will briefly summarize the regulation of renal Pi Transporters in various Pi-wasting disorders and highlight the role of a relatively new member of the Na/Pi coTransporter family: the type IIc Na/Pi Transporter/SLC34A3.
Chijiunn Pan - One of the best experts on this subject based on the ideXlab platform.
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functional analysis of mutations in the glucose 6 Phosphate Transporter that cause glycogen storage disease type ib
Molecular Genetics and Metabolism, 2008Co-Authors: Shihyin Chen, Chijiunn Pan, Soojung Lee, Wentao Peng, Janice Y. ChouAbstract:Abstract The glucose-6-Phosphate Transporter (G6PT) deficient in glycogen storage disease type Ib is a Phosphate (P i )-linked antiporter capable of G6P: P i and P i :P i exchanges. We previously characterized G6PT mutations by measuring G6P uptake activities in microsomes co-expressing G6PT and glucose-6-phosphatase-α. Here we report a new assay, based on reconstituted proteoliposomes carrying only G6PT, and characterize G6P and P i uptake activities of 23 G6PT mutations. We show that co-expression and G6PT-only assays are equivalent in measuring G6PT activity. However, the p.Q133P mutation exhibits differential G6P and P i transport activities, suggesting that characterizing G6P and P i transport activities of G6PT mutations may yield insights to this genetic disorder.
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impaired glucose homeostasis neutrophil trafficking and function in mice lacking the glucose 6 Phosphate Transporter
Human Molecular Genetics, 2003Co-Authors: Liyuan Chen, Chijiunn Pan, Jengjer Shieh, Baochuan Lin, Jiliang Gao, Philip M Murphy, Thomas F Roe, S W Moses, Jerrold M Ward, Eric J LeeAbstract:Glycogen storage disease type Ib (GSD-Ib) is caused by a deficiency in the glucose-6-Phosphate Transporter (G6PT). In addition to disrupted glucose homeostasis, GSD-Ib patients have unexplained and unexpected defects in neutrophil respiratory burst, chemotaxis and calcium flux, in response to the bacterial peptide f-Met-Leu-Phe, as well as intermittent neutropenia. We generated a G6PT knockout (G6PT-/-) mouse that mimics all known defects of the human disorder and used the model to further our understanding of the pathogenesis of GSD-Ib. We demonstrate that the neutropenia is caused directly by the loss of G6PT activity; that chemotaxis and calcium flux, induced by the chemokines KC and macrophage inflammatory protein-2, are defective in G6PT-/- neutrophils; and that local production of these chemokines and the resultant neutrophil trafficking in vivo are depressed in G6PT-/- ascites during an inflammatory response. The bone and spleen of G6PT-/- mice are developmentally delayed and accompanied by marked hypocellularity of the bone marrow, elevation of myeloid progenitor cell frequencies in both organs and a corresponding dramatic increase in granulocyte colony stimulating factor levels in both GSD-Ib mice and humans. So, in addition to transient neutropenia, a sustained defect in neutrophil trafficking due to both the resistance of neutrophils to chemotactic factors, and reduced local production of neutrophil-specific chemokines at sites of inflammation, may underlie the myeloid deficiency in GSD-Ib. These findings demonstrate that G6PT is not just a G6P transport protein but also an important immunomodulatory protein whose activities need to be addressed in treating the myeloid complications in GSD-Ib patients.
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structure function analysis of the glucose 6 Phosphate Transporter deficient in glycogen storage disease type ib
Human Molecular Genetics, 2002Co-Authors: Liyuan Chen, Chijiunn Pan, Jengjer Shieh, Janice Y. ChouAbstract: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 structure– function 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.
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inactivation of the glucose 6 Phosphate Transporter causes glycogen storage disease type 1b
Journal of Biological Chemistry, 1999Co-Authors: Hisayuki Hiraiwa, Baochuan Lin, Chijiunn Pan, S W Moses, Janice Y. ChouAbstract:Glycogen storage disease type 1b (GSD-1b) is proposed to be caused by a deficiency in microsomal glucose 6-Phosphate (G6P) transport, causing a loss of glucose-6-phosphatase activity and glucose homeostasis. However, for decades, this disorder has defied molecular characterization. In this study, we characterize the structural organization of the G6P Transporter gene and identify mutations in the gene that segregate with the GSD-1b disorder. We report the functional characterization of the recombinant G6P Transporter and demonstrate that mutations uncovered in GSD-1b patients disrupt G6P transport. Our results, for the first time, define a molecular basis for functional deficiency in GSD-1b and raise the possibility that the defective G6P Transporter contributes to neutropenia and neutrophil/monocyte dysfunctions characteristic of GSD-1b patients.
Tsuyoshi Nishi - One of the best experts on this subject based on the ideXlab platform.
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mfsd2b is a sphingosine 1 Phosphate Transporter in erythroid cells
Scientific Reports, 2018Co-Authors: Naoki Kobayashi, Shoko Kawasakinishi, Masato Otsuka, Yu Hisano, Akihito Yamaguchi, Tsuyoshi NishiAbstract:Sphingosine 1-Phosphate (S1P) is an intercellular signaling molecule present in blood. Erythrocytes have a central role in maintaining the S1P concentration in the blood stream. We previously demonstrated that S1P is exported from erythrocytes by a glyburide-sensitive S1P Transporter. However, the gene encoding the S1P Transporter in erythrocytes is unknown. In this study, we found that the mouse erythroid cell line, MEDEP-E14, has S1P export activity and exhibits properties that are consistent with those of erythrocytes. Using microarray analysis of MEDEP-E14 cells and its parental cell line, E14TG2a, we identified several candidate genes for S1P export activity. Of those genes, only one gene, Mfsd2b, showed S1P transport activity. The properties of S1P release by MFSD2B were similar to those in erythrocytes. Moreover, knockout of MFSD2B in MEDEP-E14 cells decreased S1P export from the cells. These results strongly suggest that MFSD2B is a novel S1P Transporter in erythroid cells.
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mouse spns2 functions as a sphingosine 1 Phosphate Transporter in vascular endothelial cells
PLOS ONE, 2012Co-Authors: Yu Hisano, Naoki Kobayashi, Akihito Yamaguchi, Tsuyoshi NishiAbstract:Sphingosine-1-Phosphate (S1P), a sphingolipid metabolite that is produced inside the cells, regulates a variety of physiological and pathological responses via S1P receptors (S1P1–5). Signal transduction between cells consists of three steps; the synthesis of signaling molecules, their export to the extracellular space and their recognition by receptors. An S1P concentration gradient is essential for the migration of various cell types that express S1P receptors, such as lymphocytes, pre-osteoclasts, cancer cells and endothelial cells. To maintain this concentration gradient, plasma S1P concentration must be at a higher level. However, little is known about the molecular mechanism by which S1P is supplied to extracellular environments such as blood plasma. Here, we show that SPNS2 functions as an S1P Transporter in vascular endothelial cells but not in erythrocytes and platelets. Moreover, the plasma S1P concentration of SPNS2-deficient mice was reduced to approximately 60% of wild-type, and SPNS2-deficient mice were lymphopenic. Our results demonstrate that SPNS2 is the first physiological S1P Transporter in mammals and is a key determinant of lymphocyte egress from the thymus.
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characterization of the atp dependent sphingosine 1 Phosphate Transporter in rat erythrocytes
Journal of Biological Chemistry, 2009Co-Authors: Naoki Kobayashi, Akihito Yamaguchi, Nobuyoshi Kobayashi, Tsuyoshi NishiAbstract:Sphingosine 1-Phosphate (S1P) is a bioactive lipid signal transmitter present in blood. Blood plasma S1P is supplied from erythrocytes and plays an important role in lymphocyte egress from lymphoid organs. However, the S1P export mechanism from erythrocytes to blood plasma is not well defined. To elucidate the mechanism of S1P export from erythrocytes, we performed the enzymatic characterization of S1P Transporter in rat erythrocytes. Rat erythrocytes constitutively released S1P without any stimulus. The S1P release was reduced by an ABCA1 Transporter inhibitor, glyburide, but not by a multidrug resistance-associated protein inhibitor, MK571, or a multidrug resistance protein inhibitor, cyclosporine A. Furthermore, we measured S1P transport activity using rat erythrocyte inside-out membrane vesicles (IOVs). Although the effective S1P transport into IOVs was observed in the presence of ATP, this activity was also supported by dATP and adenosine 5'-(beta,gamma-imido)triPhosphate. The rate of S1P transport increased depending on S1P concentration, with an apparent K(m) value of 21 microm. Two phosphorylated sphingolipids, dihydrosphingosine 1-Phosphate and ceramide 1-Phosphate, did not inhibit S1P transport. Similar to the intact erythrocytes, the uptake of S1P into IOVs was inhibited by glyburide and vanadate but not by the other ABC Transporter inhibitors. These results suggest that S1P is exported from the erythrocytes by a novel ATP-dependent Transporter.