ABCMdb

PMID: 19506252

Woodward OM, Kottgen A, Coresh J, Boerwinkle E, Guggino WB, Kottgen M
Identification of a urate transporter, ABCG2, with a common functional polymorphism causing gout.
Proc Natl Acad Sci U S A. 2009 Jun 23;106(25):10338-42. Epub 2009 Jun 8., 2009-06-23 [PubMed]

Sentences

No. Mutations Sentence Comment

4 ABCG2 p.Gln141Lys Introduction of the mutation Q141K encoded by the common SNP rs2231142 by site-directed mutagenesis resulted in 53% reduced urate transport rates compared to wild-type ABCG2 (P < 0.001). Login to comment 35 ABCG2 p.Gln141Lys Impaired Urate Transport of ABCG2 Q141K. Login to comment 47 ABCG2 p.Gln141Lys To test whether this SNP is not only statistically associated but causally related to elevated urate levels, we introduced the mutation Q141K encoded by the rs2231142 T allele by site-directed mutagenesis. Login to comment 48 ABCG2 p.Gln141Lys Wild-type and Q141K mutant ABCG2 showed similar expression levels at the cell surface when expressed in Xenopus oocytes (Fig. S1). Login to comment 49 ABCG2 p.Gln141Lys Notably, Q141K-expressing oocytes showed 54% reduced urate transport rates compared with those expressing wild-type ABCG2 at similar levels (Fig. 2 B and C), and decreased urate efflux across a range of intracellular urate concentrations (Fig. 2D). Login to comment 50 ABCG2 p.Gln141Lys Our evidence for Q141K as a causal loss-of-function variant is supported by data showing reduced transport of chemotherapeutic agents by this mutation (6). Login to comment 53 ABCG2 p.Gln141Lys Information on the SNP rs2231442, encoding the Q141K variant, was available as an imputed SNP from genome-wide Affymetrix 6.0 data along with 601 other SNPs in a 600-kb region containing ABCG2, and also genotyped directly using the TaqMan assay. Login to comment 64 ABCG2 p.Gln141Lys Among the white study participants, population-attributable risk calculations yielded that a conservatively estimated 10% of gout cases could be attributed to the Q141K mutation. Login to comment 65 ABCG2 p.Gln141Lys Our findings are of substantial clinical interest because of the high prevalence of the Q141K mutation in individuals of European and Asian ancestry and the substantial population attributable risk. Login to comment 69 ABCG2 p.Ser187Thr The physiological importance of ABCG2 in humans is illustrated by 0.0 0.5 1.0 1.5 noitalumuccAetarU nim021/etycoo/lomp H2O WT WT +FTC S187T 0 100 200 300 400 0 2 4 noitalumuccAetarU nim06/etycoo/lomp Urate Concentration ( µM) 0 2 4 6 8 10 0.00 0.05 0.10 nim/etycoo/lomp:xulffE Internal oocyte concentration (µM) 0 20 40 60 0.4 0.6 0.8 1.0 gniniameretarUevitaleR * 0.0 0.5 1.0 1.5 noitalumuccAetarU nim021/etycoo/lomp H2O MRP4ABCG2 ** A B C D E ** ** 500 ** ** ** ** * ** ** ** G F Time (min) 0.0 0.5 1.0 1.5 2.0 eAccumulationtarUevitaleR ** LLCPK : control +FTC basal brush border membrane Fig. 1. Login to comment 72 ABCG2 p.Ser187Thr (B) Urate accumulation in oocytes injected with H2O, ABCG2 WT mRNA (incubated with or without 5 ␮M FTC), or the nonfunctional ABCG2 mutant S187T (N ϭ 10 samples each and n ϭ 20 oocytes each for all accumulation experiments). Login to comment 73 ABCG2 p.Ser187Thr (C) Urate accumulation is dependent on the extracellular urate concentration (H2O [blue circles]; S187T [black squares]; WT [red triangles]; N ϭ 10 and n ϭ 20 each). Login to comment 82 ABCG2 p.Gln141Lys We show that ABCG2 is a urate efflux transporter and rs2231142 (Q141K) a loss-of-function mutation that causes hyperuricemia and gout. Login to comment 89 ABCG2 p.Gln141Lys Oocytes after injection were cultured in a modified L-15 media (OR-3) and kept at 15-20 °C. mRNA was prepared using the SP6 mMessage mMachine (Ambion Inc.) according to the manufactures protocol. The Q141K mutant was created using site directed mutagenesis with the primers: 5Ј-GGT GAG AGA AAA CTT AAA GTT CTC AGC AGC TC 3-Ј and 5Ј-GAG CTG CTG AGA ACT TTA AGT TTT CTC TCA CC-3Ј and completed using the Quick-Change Lightning kit (Stratagene) according to the manufacturer`s protocol. Login to comment 97 ABCG2 p.Gln141Lys For the 1-2-h accumulation studies, after incubation in the C-14 urate solution, 2 oocytes were placed in each scintillation tube (N) with lysis buffer of 1 N NaOH; a total Q141 *CFTR F508 ABCG2 CFTR 1 2 3 4 5 0.02 0.04 0.06 0.08 0.10 Internal oocyte concentration ( µM) nim/etycoo/lomp:xulffE 0.0 0.3 0.6 0.9 noitalumuccAetarU nim021/etycoo/lomp WT Q141K C D** 150 76 52 H2OWTQ141K α-BCRP α-actin B A Fig. 2. Login to comment 98 ABCG2 p.Gln141Lys The ABCG2 Q141K mutation results in reduced urate transport. Login to comment 102 ABCG2 p.Gln141Lys (B) Western blot of oocytes expressing ABCG2 WT or Q141K (monomers and dimers of ABCG2 are detected). Login to comment 104 ABCG2 p.Gln141Lys (C) Accumulation rates in oocytes expressing ABCG2 WT or Q141K normalized to ABCG2 expression level. Login to comment 105 ABCG2 p.Gln141Lys (D) Efflux rates for ABCG2 WT and Q141K. Login to comment 106 ABCG2 p.Gln141Lys (WT [red triangles]: slope ϭ 0.01, N ϭ 55; Q141K [black squares]: slope ϭ 0.02, N ϭ 55). Login to comment