Mutations In SLC22A12 Research Articles

URAT1 and GLUT9 mutations in Spanish patients with renal hypouricemia

BackgroundRenal hypouricemia (RHUC), a rare inherited disorder characterized by impaired uric acid (UA) reabsorption in the proximal tubule, is caused by mutations in SLC22A12 or SLC2A9. Most mutations have been identified in Japanese patients, and only a few have been detected in Europeans. MethodsWe report clinical, biochemical and genetics findings of fourteen Spanish patients, six Caucasians and eight of Roma ethnia, diagnosed with idiopathic RHUC. Two of the patients presented exercise-induced acute renal failure and another one had several episodes of nephrolithiasis and four of them had progressive deterioration of renal function, while the rest were asymptomatic. ResultsMolecular analysis revealed SLC22A12 mutations in ten of the patients, and SLC2A9 mutations in the other four. A new heterozygous SLC22A12 missense mutation, c.1427C>A (p.A476D), was identified in two affected members of the same family. The rest of the patients presented homozygous, heterozygous or compound heterozygous mutations that have been previously identified in patients with RHUC; SLC22A12 p.T467M and p.L415_G417del, and SLC2A9 p.T125M. Expression studies in Xenopus oocytes revealed that c.1427C>A reduced UA transport but did not alter the location of URAT1 protein on the plasma membrane. ConclusionsThe biochemical and clinical features of our patients together with the genetic analysis results confirmed the diagnosis of RHUC. This is the first report describing SLC22A12 and SLC2A9 mutations in Spanish patients.

AB0832 Prevalence and Possible Causes of Hypouricemia at A Tertiary Care Hospital in South Korea

BackgroundHypouricemia is defined as a serum uric acid concentration of less than 2.0 mg/dL and is characterized by increased uric acid clearance or decreased uric acid production. Causes of hypouricemia...

Molecular Background of Urate Transporter Genes in Patients with Exercise-Induced Acute Kidney Injury

Background/Aims: Exercise-induced acute renal failure [exercise-induced acute kidney injury (EI-AKI)] is defined as AKI due to heavy anaerobic exercise. Although hypouricemia is known to be a risk factor for the onset of EI-AKI, a direct causal link between EI-AKI and serum uric acid has not been established. This study aimed to analyze urate transporter genes in patients with EI-AKI and its molecular mechanism. Methods: Genomic DNA and total RNA were isolated from peripheral blood leukocytes of patients with a history of EI-AKI. Mutations were analyzed by PCR and a direct sequencing method. We first analyzed the SLC22A12 gene, and then the SLC2A9 gene if no mutations were found in SLC22A12. Results: Seventeen patients were enrolled in this study and 16 had mutations: 15 in SLC22A12 and 1 in SLC2A9. Fourteen (82.4%) patients showed hypouricemia, and all of the patients with hypouricemia had either homozygous or compound heterozygous mutations in SLC22A12 or SLC2A9, which confirmed that all of them had renal hypouricemia. Two patients had heterozygous mutations of SLC22A12, and they were not accompanied by hypouricemia. One patient was found to have no mutations in SLC22A12 or SLC2A9. Conclusion: We were able to determine the genetic background of urate transporter genes in patients with EI-AKI. Decreased function of urate transporters, rather than decreased serum uric acid levels, may be of great importance for the onset of EI-AKI.

Simple and rapid detection method for the mutations in SLC22A12 that cause hypouricemia by allele-specific real-time polymerase chain reaction

BackgroundHypouricemia is a disorder that serum urate level is less than 2.0mg/dl, and relatively common in the Japanese population, where the main genetic cause of hypouricemia is W258X and R90H mutations in human urate trasnsporter 1(SLC22A12). Small scale screening has relied on time-consuming traditional ways like polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Therefore, it is beneficial that we have an easy and rapid detection method for these mutations. MethodsIn this report, we established a touchdown allele-specific real-time polymerase chain reaction (ASPCR) assay for detecting W258X and R90H mutations in SLC22A12, respectively. ResultsQuantifiable discrimination was successfully achieved by ∆Ct value. Furthermore, we conducted W258X and R90H screening against 120 control genome sets, whereby frequency was 2.92% for W258X, and not detected for R90H, respectively. ConclusionsThe two mutations, W258X and R90H in SLC22A12 were successfully genotyped by an easy and rapid ASPCR assay.

Clinical and Functional Characterization of URAT1 Variants

Idiopathic renal hypouricaemia is an inherited form of hypouricaemia, associated with abnormal renal handling of uric acid. There is excessive urinary wasting of uric acid resulting in hypouricaemia. Patients may be asymptomatic, but the persistent urinary abnormalities may manifest as renal stone disease, and hypouricaemia may manifest as exercise induced acute kidney injury. Here we have identified Macedonian and British patients with hypouricaemia, who presented with a variety of renal symptoms and signs including renal stone disease, hematuria, pyelonephritis and nephrocalcinosis. We have identified heterozygous missense mutations in SLC22A12 encoding the urate transporter protein URAT1 and correlate these genetic findings with functional characterization. Urate handling was determined using uptake experiments in HEK293 cells. This data highlights the importance of the URAT1 renal urate transporter in determining serum urate concentrations and the clinical phenotypes, including nephrolithiasis, that should prompt the clinician to suspect an inherited form of renal hypouricaemia.

Recurrent EIARF and PRES With Severe Renal Hypouricemia by Compound Heterozygous SLC2A9 Mutation

Renal hypouricemia (RHU) is a hereditary disease that predisposes affected people to exercise-induced acute renal failure (EIARF). In most patients with RHU, the disorder is caused by loss-of-function mutations in SLC22A12 (solute carrier family 22, member 12), which encodes urate transporter 1 (URAT1). Patients with RHU without any mutations in the URAT1 gene were recently found to have a mutation in the glucose transporter 9 (GLUT9) gene (SLC2A9 [solute carrier family 2, member 9]). Central nervous system complications seem to be rare in patients with RHU with SLC22A12 mutations. Here, we report the case of a girl with severe RHU (serum urate: 5.9 μmol/L [0.1 mg/dL]) associated with recurrent EIARF in whom the disease was caused by a compound heterozygous mutation in SLC2A9, a nonsense mutation in the paternal allele (p.G207X in exon 7), and a large duplication (c.1-2981_1204+16502) in the maternal allele detected by reverse-transcription polymerase chain reaction (PCR), semiquantitative PCR, long PCR, and direct sequencing. The episodes of EIARF were complicated by posterior reversible encephalopathy syndrome (PRES), which suggested a relationship between PRES and GLUT9 or severe hypouricemia. This is the second report of mutations of both alleles of SLC2A9 that resulted in severe hypouricemia. Our findings indicate that even a nonsense mutation responsible for the heterozygous status of SLC2A9 did not cause severe hypouricemia, and they lend support to previous speculation that mutations of both SLC2A9 alleles cause severe hypouricemia. Our case shows that GLUT9, unlike URAT1, may play a specific role in exercise-induced PRES.

Genetic Analysis of Human Urate Transporter 1 (hURAT1) and Glucose Transporter 9 (GLUT9) in Taiwanese Patients with Idiopathic Renal Hypouricemia

Background: The phenotypic characteristics of idiopathic renal hypouricemia (IRH) include impaired renal uric acid re-absorption, low serum uric acid levels, and severe complications such as exercise-induced acute renal failure (EIARF) and nephrolithiasis. IRH is increasingly reported to be associated with inactivating mutations in either SLC22A12 gene encoding the human urate acid transporter 1 (hURAT1) or 5LC2A9 gene encoding glucose transporter 9 (GLUT9) in proximal tubules. Mutations in hURAT1 or GLUT9 have not been reported in Taiwanese patients with IRH. Purpose: To investigate the phenotypic and genetic characteristics in Taiwanese patients with IRH. Patients: Six Taiwanese patients (5 males and 1 female, mean age 30±23 years) with hypouricemia (serum uric acid concentration＜2.3 mg/dL) and concomitant excessive renal uric acid excretion (FE(subscript UA)＞10%) from five unrelated families were enrolled. Their clinical symptoms and biochemical studies were recorded. Methods: Molecular analysis of both SLC22A12 and 5LC2A9 genes was performed by polymerase chain reaction (PCR)-single strand conformation polymorphism (SSCP) and direct sequencing. Results: Serum uric acid concentration was 1.16±0.23 mg/dl with a fractional excretion of uric acid of 61±12%. Two male patients experienced exercise-induced acute renal failure requiring hemodialysis. None of them had nephrocalcinosis or uric acid stone. Despite a negative mutation in 5LC2J49, four novel SLC22A12 mutations (W120R, M2 15L, T2 17M and T467M) and one recurrent homozygous (R90H) mutations were identified in five patients. Conclusion: EIARF can develop in Taiwanese patients with IRH, and most of them are males. Mutations in 5LC22A 12 rather than 5LC2A9 cause IRH in patients with FE(subscript UA)＜100%. Other candidate genes need to be identified for IRH patients without 5LC22J4 12 and SLC2A9 gene mutations.

Functional characterization of a novel urate efflux transporter URATv1 (SLC2A9) and its relation to renal hypouricemia.

Reanl urate handling is clinically important because underexcretion of urate causes hyperuricemia. In 2002, we identified an apical urate‐anion exchanger, URAT1 encoded by SLC22A12. But it is currently not known how urate taken up via URAT1 exits from the tubular cell. In this study, we functionally characterized SLC2A9 gene product GLUT9 in search of urate excreting transporter. GLUT9 expressed Xenopus oocytes transported [14C]urate time‐ and concentration‐dependent manner (Km, 365 µM). Interestingly, we found that its transport was Na+‐independent and enhanced at high concentrations of extracellular potassium, indicating that GLUT9‐mediated urate transport is affected by the changes in membrane potential. GLUT9‐expressing oocytes preloaded with radiolabelled urate showed a time‐dependent efflux of radioactivity in the standard uptake solution Thus, we renamed this novel urate transporter as voltage‐driven urate transporter (URATv1). In vivo role of URATv1 is supported by the fact that a renal hypouricemia patient without any mutations in SLC22A12 was found to have a missense mutation P412R in SLC2A9, which reduced urate transport activity in vitro. Considering its basolateral expression in proximal tubules in human kidney, URATv1 is proposed to be a basolateral exit pathway of urate, which is likely to act in tandem with URAT1 for urate reabsorption.

Prevalence of hypouricaemia and SLC22A12 mutations in healthy Korean subjects

Mutations in the SLC22A12 gene, which encodes a uric acid transporter, URAT1, are associated with renal hypouricaemia. This study was designed to measure serum uric acid (Sua) levels and allele frequencies of two common mutations in SLC22A12, W258X and R90H, in healthy Korean subjects. A total of 909 unrelated Korean adults (male : female, 1:1.23; mean age, 48.4 +/- 11.0 years) were recruited among those who had taken a routine health check-up in a health centre in 2003. None of them had hypertension, diabetes mellitus, kidney diseases or liver diseases. Genotyping for W258X and R90H was performed using the TaqMan method. The prevalences of hyperuricaemia (Sua levels, >416 micromol/L) and hypouricaemia (Sua levels, <178 micromol/L) were 4.6% and 3.3%, respectively. A marked male preponderance in the hyperuricaemic group was noted, and the men revealed higher Sua than the women. The Sua showed a positive correlation with serum creatinine level and blood pressure. In the hypouricaemic group, the allele frequencies of W258X and R90H were 11.7% and 6.7%, respectively, and the proportion of subjects with one or both of the mutant alleles was 33.3%. Hyperuricaemic subjects never had either mutation. The W258X and/or R90H mutations in the SLC22A12 gene are one of the major factors responsible for hypouricaemia, and one-third of the hypouricaemic subjects had one or both of the mutant alleles.

Plasma Urate Level Is Directly Regulated by a Voltage-driven Urate Efflux Transporter URATv1 (SLC2A9) in Humans

Hyperuricemia is a significant factor in a variety of diseases, including gout and cardiovascular diseases. Although renal excretion largely determines plasma urate concentration, the molecular mechanism of renal urate handling remains elusive. Previously, we identified a major urate reabsorptive transporter, URAT1 (SLC22A12), on the apical side of the renal proximal tubular cells. However, it is not known how urate taken up by URAT1 exits from the tubular cell to the systemic circulation. Here, we report that a sugar transport facilitator family member protein GLUT9 (SLC2A9) functions as an efflux transporter of urate from the tubular cell. GLUT9-expressed Xenopus oocytes mediated saturable urate transport (K(m): 365+/-42 microm). The transport was Na(+)-independent and enhanced at high concentrations of extracellular potassium favoring negative to positive potential direction. Substrate specificity and pyrazinoate sensitivity of GLUT9 was distinct from those of URAT1. The in vivo role of GLUT9 is supported by the fact that a renal hypouricemia patient without any mutations in SLC22A12 was found to have a missense mutation in SLC2A9, which reduced urate transport activity in vitro. Based on these data, we propose a novel model of transcellular urate transport in the kidney; urate [corrected] is taken up via apically located URAT1 and exits the cell via basolaterally located GLUT9, which we suggest be renamed URATv1 (voltage-driven urate transporter 1).

Molecular analysis of the SLC22A12 (URAT1) gene in patients with primary gout

To analyse the SLC22A12 (URAT1) gene in primary gout patients, first-grade relatives and healthy controls and the possible association of them with demographic and clinical data. We included 69 consecutive patients with diagnosis of primary gout, as well as 29 first-grade relatives and 120 healthy volunteers. Demographic and clinical data were obtained from the patients and relatives. DNA was purified from peripheral blood and all 10 exons of the SLC22A12 (URAT1) gene were sequenced. We found six different mutations in the SLC22A12 gene in 16 out of 69 (23%) patients with primary gout. Five mutations were in exon 5 and one in exon 4; five out of six mutations were heterozygous (one compound heterozygous) and one homozygous. The C850G mutation (exon 5) was found in 11 gout patients, these patients have lower levels of triglycerides than the rest of the group: 160 +/- 56 vs 292 +/- 203 mg/dl (P = 0.038). In one family, we found SLC22A12 mutations in three relatives within exon 5. We did not find mutations in the other exons studied (1-3 and 6-10), nor in any of the 10 exons of the 120 healthy volunteers. We found several mutations in SLC22A12 gene associated with primary gout, the definite role of these mutations in URAT1 activity needs to be further studied.

Mutational analysis of idiopathic renal hypouricemia in Korea

Idiopathic renal hypouricemia is a hereditary disease characterized by abnormally high renal uric acid clearance. Most patients are clinically silent, but acute renal failure (ARF), urolithiasis, or hematuria may develop. A defect in the SLC22A12 gene, which encodes the renal uric acid transporter, URAT1, is the known major cause of this disorder. We performed a mutational analysis of the SLC22A12 gene in five Korean patients with idiopathic renal hypouricemia in this study. Two patients presented with microscopic hematuria, one with uric acid urolithiasis, and one with exercise-induced ARF. One patient was asymptomatic. Three different mutations, W258X, R90H and R477H, were detected in four of the patients. However, no mutation was found in the fifth ARF patient. This is the first study of SLC22A12 mutations in a country other than Japan. W258X was found to be the predominant SLC22A12 mutation in Korean renal hypouricemia patients, as has been reported in Japan.

The W258X mutation in SLC22A12 is the predominant cause of Japanese renal hypouricemia.

Recently, a urate transporter, hURAT1 (human uric acid transporter 1) encoded by SLC22A12, was isolated from the human kidney. hURAT1 is presumed to play the central role in reabsorption of urate from glomerular filtrate. In the present study, we analyzed SLC22A12 in seven unrelated Japanese patients with renal hypouricemia whose serum level of urate was less than 1.0 mg/dl, and their family members. We performed direct DNA sequencing of the exon and exon-intron boundaries of SLC22A12 using genomic DNA. Six of the seven patients (86%) possess mutations in SLC22A12. In five patients, a homozygous G to A transition at nucleotide 774 within exon 4 of SLC22A12, which forms a stop codon (TGA) at codon 258 (TGG), was identified (W258X). In one patient, the C to T transition within exon 3, which changes threonine at codon 217 to methionine (T217 M), and the W258X mutation were found (compound heterozygote). Thus, among 12 mutational alleles in six patients, 11 were the W258 X mutation (92%). Family members with the heterozygous W258X mutation (carriers) show relatively low levels of serum urate. The present study demonstrates that homozygous W258X mutation is the predominant genetic cause of idiopathic renal hypouricemia in Japanese patients.

Clinical and molecular analysis of patients with renal hypouricemia in Japan-influence of URAT1 gene on urinary urate excretion.

Renal hypouricemia is an inherited and heterogeneous disorder characterized by increased urate clearance (CUA). The authors recently established that urate was reabsorbed via URAT1 on the tubular apical membrane and that mutations in SLC22A12 encoding URAT1 cause renal hypouricemia. This study was undertaken to elucidate and correlate clinical and genetic features of renal hypouricemia. The SLC22A12 gene was sequenced in 32 unrelated idiopathic renal hypouricemia patients, and the relationships of serum urate levels, and CUA/creatinine clearance (Ccr) to SLC22A12 genotype were examined. Uricosuric (probenecid and benzbromarone) and anti-uricosuric drug (pyrazinamide) loading tests were also performed in some patients. Three patients had exercise-induced acute renal failure (9.4%), and four patients had urolithiasis (12.5%). The authors identified eight new mutations and two previously reported mutations that result in loss of function. Thirty patients had SLC22A12 mutations; 24 homozygotes and compound heterozygotes, and 6 heterozygotes. Mutation G774A dominated SLC22A12 mutations (74.1% in 54 alleles). Serum urate levels were significantly lower and CUA/Ccr was significantly higher in heterozygotes compared with healthy subjects; these changes were even more significant in homozygotes and compound heterozygotes. These CUA/Ccr relations demonstrated a gene dosage effect that corresponds with the difference in serum urate levels. In contrast to healthy subjects, the CUA/Ccr of patients with homozygous and compound heterozygous SLC22A12 mutations was unaffected by pyrazinamide, benzbromarone, and probenecid. The findings indicate that SLC22A12 was responsible for most renal hypouricemia and that URAT1 is the primary reabsorptive urate transporter, targeted by pyrazinamide, benzbromarone, and probenecid in vivo.