Transcriptome-based reconstructions from the murine knockout suggest involvement of the urate transporter, URAT1 (slc22a12), in novel metabolic pathways

Abstract

URAT1 (slc22a12) was identified as the transporter responsible for renal reabsorption of the medically important compound, uric acid. However, subsequent studies have indicated that other transporters make contributions to this process, and that URAT1 transports other organic anions besides urate (including several in common with the closely related multi-specific renal organic anion transporters, OAT1 (slc22a6) and OAT3 (slc22a8)). These findings raise the possibility that urate transport is not the sole physiological function of URAT1. We previously characterized mice null for the murine ortholog of URAT1 (mURAT1; previously cloned as RST), finding a relatively modest decrement in urate reabsorptive capacity. Nevertheless, there were shifts in the plasma and urinary concentrations of multiple small molecules, suggesting significant metabolic changes in the knockouts. Although these molecules remain unidentified, here we have computationally delineated the biochemical networks consistent with transcriptomic data from the null mice. These analyses suggest alterations in the handling of not only urate but also other putative URAT1 substrates comprising intermediates in nucleotide, carbohydrate, and steroid metabolism. Moreover, the analyses indicate changes in multiple other pathways, including those relating to the metabolism of glycosaminoglycans, methionine, and coenzyme A, possibly reflecting downstream effects of URAT1 loss. Taken together with the available substrate and metabolomic data for the other OATs, our findings suggest that the transport and biochemical functions of URAT1 overlap those of OAT1 and OAT3, and could contribute to our understanding of the relationship between uric acid and the various metabolic disorders to which it has been linked.

Keywords: URAT1 (slc22a12); metabolic networks; organic anion transporter; transcriptomics; urate.

Fig. 1

Expression in mURAT1 knockout mouse kidneys of urate-regulating genes. Gene expression levels in mURAT1 knockout (KO) and wild-type (WT) mice were determined using microarrays (A) or quantitative PCR (B) and were normalized for each gene to the mean level of expression in the WT. XDH, xanthine dehydrogenase; HGPRT, hypoxanthine guanine phosphoribosyl transferase; ADA, adenosine deaminase; AMPD2, adenosine monophosphate deaminase 2; PNP, purine nucleoside phosphorylase; UAT, urate transporter (galectin 9); MRP2, multidrug resistance protein 2 (ATP-Binding Cassette Sub-Family C Member 2); MRP4, multidrug resistance protein 4 (ATP-Binding Cassette, Sub-Family C, Member 4); ABCG2, ATP-binding cassette, sub-family G, member 2; NPT1, sodium-phosphate transport protein 1 (slc17a1); NPT4, sodium-phosphate transport protein 4 (slc17a3); GLUT9, glucose transporter type 9 (slc2a9); OAT10, organic anion transporter 10 (organic cation transporter like 3; slc22a13); OAT1, organic anion transporter 1 (slc22a6); OAT3, organic anion transporter 3 (slc22a8). Data in Panel A for XDH, HGPRT, ADA, AMPD2, PNP, UAT, MRP2, MRP4, NPT1, OAT1, OAT3, and mURAT1 were reported previously . Values represent mean±standard error.

Fig. 2

Gene ontology functional annotation enrichment among genes differentially expressed or variable in the mURAT1 knockout. The enrichment of gene ontology (GO) functional annotation terms among genes that are differentially expressed (panel A) or differentially variable (panel B) in the mURAT1 knockout compared with wild-type mice was determined using AmiGO. GO terms are depicted in boxes shaded according to the statistical significance of their enrichment and p values <10⁻⁵ are indicated.

Fig. 3

Metabolic sub-systems containing reactions with two-fold or greater change in flux-span in mURAT1 knockout reconstructions. Proportions of the indicated metabolic sub-systems comprising reactions having flux-spans increased (panel A) or decreased (panel B) by two-fold or greater in the mURAT1 knockout relative to wild-type reconstructions are depicted. The actual number of reactions increased or decreased by two-fold or greater than is indicated above each column. Error bars denote the standard error of the proportion; black columns, p<0.001; dark gray columns, p<0.01; light gray columns, p<0.05; open columns, NS.