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Objective: Using European descent Czech populations, we performed a study of SLC2A9 and SLC22A12 genes previously identified as being associated with serum uric acid concentrations and gout. This is the first study of the impact of non-synonymous allelic variants on the function of GLUT9 except for patients suffering from renal hypouricemia type 2., Methods: The cohort consisted of 250 individuals (150 controls, 54 nonspecific hyperuricemics and 46 primary gout and/or hyperuricemia subjects). We analyzed 13 exons of SLC2A9 (GLUT9 variant 1 and GLUT9 variant 2) and 10 exons of SLC22A12 by PCR amplification and sequenced directly. Allelic variants were prepared and their urate uptake and subcellular localization were studied by Xenopus oocytes expression system. The functional studies were analyzed using the non-parametric Wilcoxon and Kruskall-Wallis tests; the association study used the Fisher exact test and linear regression approach., Results: We identified a total of 52 sequence variants (12 unpublished). Eight non-synonymous allelic variants were found only in SLC2A9: rs6820230, rs2276961, rs144196049, rs112404957, rs73225891, rs16890979, rs3733591 and rs2280205. None of these variants showed any significant difference in the expression of GLUT9 and in urate transport. In the association study, eight variants showed a possible association with hyperuricemia. However, seven of these were in introns and the one exon located variant, rs7932775, did not show a statistically significant association with serum uric acid concentration., Conclusion: Our results did not confirm any effect of SLC22A12 and SLC2A9 variants on serum uric acid concentration. Our complex approach using association analysis together with functional and immunohistochemical characterization of non-synonymous allelic variants did not show any influence on expression, subcellular localization and urate uptake of GLUT9.

Renal hypouricemia (RHUC) is a heterogeneous inherited disorder characterized by impaired tubular uric acid (UA) transport with severe complications, such as acute kidney injury (AKI). Type 1 is caused by a loss-of-function mutation in the SLC22A12 gene (URAT1), type 2 in the SLC2A9 gene (GLUT9). This article describes three Czech families with RHUC type 1. The serum UA in the probands was 0.9, 1.1 and 0.5 mg/dl and expressed as an increase in the fractional excretion of UA (48, 43 and 39%). The sequencing analysis of SLC22A12 revealed three novel variants: p.G366R, p.T467M and a deletion p.L415_G417del. A detailed metabolic investigation in proband C for progressive visual failure supported suspicion of neuronal ceroid lipofuscinosis type 7 conditioned by the mutation in the MFSD8 gene. Functional studies showed significantly decreased urate uptake and a mis-localized URAT1 signal in p.G366R, p.L415_G417del and p.T467M. Furthermore, colocalization studies showed accumulation of URAT1 protein in the endoplasmic reticulum. The findings suggest that loss-of-function mutations cause RHUC via loss of UA absorption partly by protein misfolding. However, they do not necessarily lead to AKI and a possible genotype-phenotype correlation was not proposed. Furthermore, results confirm an uneven geographical and ethnic distribution of SLC22A12 variants; the p.L415_G417del mutation predominates in the Roma ethnic group in the Czech Republic.