Date Published: October 7, 2008
Publisher: Public Library of Science
Author(s): Mark J Caulfield, Patricia B Munroe, Deb O’Neill, Kate Witkowska, Fadi J Charchar, Manuel Doblado, Sarah Evans, Susana Eyheramendy, Abiodun Onipinla, Philip Howard, Sue Shaw-Hawkins, Richard J Dobson, Chris Wallace, Stephen J Newhouse, Morris Brown, John M Connell, Anna Dominiczak, Martin Farrall, G. Mark Lathrop, Nilesh J Samani, Meena Kumari, Michael Marmot, Eric Brunner, John Chambers, Paul Elliott, Jaspal Kooner, Maris Laan, Elin Org, Gudrun Veldre, Margus Viigimaa, Francesco P Cappuccio, Chen Ji, Roberto Iacone, Pasquale Strazzullo, Kelle H Moley, Chris Cheeseman, Andrew Hattersley
Abstract: BackgroundSerum uric acid levels in humans are influenced by diet, cellular breakdown, and renal elimination, and correlate with blood pressure, metabolic syndrome, diabetes, gout, and cardiovascular disease. Recent genome-wide association scans have found common genetic variants of SLC2A9 to be associated with increased serum urate level and gout. The SLC2A9 gene encodes a facilitative glucose transporter, and it has two splice variants that are highly expressed in the proximal nephron, a key site for urate handling in the kidney. We investigated whether SLC2A9 is a functional urate transporter that contributes to the longstanding association between urate and blood pressure in man.Methods and FindingsWe expressed both SLC2A9 splice variants in Xenopus laevis oocytes and found both isoforms mediate rapid urate fluxes at concentration ranges similar to physiological serum levels (200–500 μM). Because SLC2A9 is a known facilitative glucose transporter, we also tested whether glucose or fructose influenced urate transport. We found that urate is transported by SLC2A9 at rates 45- to 60-fold faster than glucose, and demonstrated that SLC2A9-mediated urate transport is facilitated by glucose and, to a lesser extent, fructose. In addition, transport is inhibited by the uricosuric benzbromarone in a dose-dependent manner (Ki = 27 μM). Furthermore, we found urate uptake was at least 2-fold greater in human embryonic kidney (HEK) cells overexpressing SLC2A9 splice variants than nontransfected kidney cells. To confirm that our findings were due to SLC2A9, and not another urate transporter, we showed that urate transport was diminished by SLC2A9-targeted siRNA in a second mammalian cell line. In a cohort of men we showed that genetic variants of SLC2A9 are associated with reduced urinary urate clearance, which fits with common variation at SLC2A9 leading to increased serum urate. We found no evidence of association with hypertension (odds ratio 0.98, 95% confidence interval [CI] 0.9 to 1.05, p > 0.33) by meta-analysis of an SLC2A9 variant in six case–control studies including 11,897 participants. In a separate meta-analysis of four population studies including 11,629 participants we found no association of SLC2A9 with systolic (effect size −0.12 mm Hg, 95% CI −0.68 to 0.43, p = 0.664) or diastolic blood pressure (effect size −0.03 mm Hg, 95% CI −0.39 to 0.31, p = 0.82).ConclusionsThis study provides evidence that SLC2A9 splice variants act as high-capacity urate transporters and is one of the first functional characterisations of findings from genome-wide association scans. We did not find an association of the SLC2A9 gene with blood pressure in this study. Our findings suggest potential pathogenic mechanisms that could offer a new drug target for gout.
Partial Text: Elevated serum urate levels are associated with important common disorders such as gout, metabolic syndrome, diabetes, hypertension, and cardiovascular morbidity and mortality [1–4]. Uric acid is principally derived from the breakdown of dietary and cellular purines. Humans and great apes are exposed to higher urate levels than other mammalian species because of the inactivation of hepatic uricase . In humans the kidney has a pivotal role in urate handling, with secretory mechanisms balanced against efficient reabsorption resulting in only 10% of the filtered load actually being excreted in the urine . The established urate transporter systems in the proximal nephron includes; the urate anion transporter (URAT1), which is a target of uricosuric drugs, multiple organic anion transporters (OATs 1–4), the urate transporter (UAT), and a voltage dependent organic anion transporter (OATv1) .
Recent genome-wide association scans identified and replicated association between SNPs at the SLC2A9 gene locus and serum urate and with gout [6,7,25,26]. In this study we have shown that both human SLC2A9 splice variants, SLC2A9a and SLC2A9b, can mediate urate fluxes at a very high rate and significantly faster than their facilitated transport of either glucose or fructose. The kinetics indicate that the transporter’s apparent capacity for substrate, or Km value (∼ 1 mM), is above the basal, physiologic plasma concentrations of urate. Moreover, the Vmax value indicates a high-capacity transporter. All of these data suggest that this membrane protein plays an important role in the handling of urate in the proximal nephron, which completely fits with the findings from genome-wide scans of common allelic variation elevating urate by 20 μmol/l per allele . In the context of everyday clinical practice this genetic influence on urate is equivalent to 5%–10% of the normal range of serum urate (180 μmol/l to 420 μmol/l), which is not trivial. These findings are confirmed by complementary functional studies on SLC2A9 in relation to urate handling and gout [25–27].