Yoon J et al. (2021), Characterization of a Compound Heterozygous SLC...

XB-ART-58488

Biomedicines 2021 Sep 06;99:. doi: 10.3390/biomedicines9091172.

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Characterization of a Compound Heterozygous SLC2A9 Mutation That Causes Hypouricemia.

Yoon J , Cachau R , David VA , Thompson M , Jung W , Jee SH , Daar IO , Winkler CA , Cho SK .

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Renal hypouricemia is a rare genetic disorder. Hypouricemia can present as renal stones or exercise-induced acute renal failure, but most cases are asymptomatic. Our previous study showed that two recessive variants of SLC22A12 (p.Trp258*, pArg90His) were identified in 90% of the hypouricemia patients from two independent cohorts: the Korean genome and epidemiology study (KoGES) and the Korean Cancer Prevention Study (KCPS-II). In this work, we investigate the genetic causes of hypouricemia in the rest of the 10% of unsolved cases. We found a novel non-synonymous mutation of SLC2A9 (voltage-sensitive uric acid transporter) in the whole-exome sequencing (WES) results. Molecular dynamics prediction suggests that the novel mutation p.Met126Val in SLCA9b (p.Met155Val in SLC2A9a) hinders uric acid transport through a defect of the outward open geometry. Molecular analysis using Xenopus oocytes confirmed that the p.Met126Val mutation significantly reduced uric acid transport but does not affect the SLC2A9 protein expression level. Our results will shed light on a better understanding of SLC2A9-mediated uric acid transport and the development of a uric acid-lowering agent.

???displayArticle.pubmedLink??? 34572357
???displayArticle.pmcLink??? PMC8471325
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Genes referenced: asb12 asic3 dnai1 gpr101 pik3cb ppef1 pwwp2b rbm12 rlim slc2a9

???displayArticle.disOnts??? urolithiasis [+]
???displayArticle.omims??? HYPOURICEMIA, RENAL, 1; RHUC1 [+]

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	Figure 1. SLC2A9b model structure overview.
	Figure 2. Mechanistic interpretation of the effects of the M126V mutation of SLC2A9b. The M126V model suggests that this mutation renders the vestibular regions unavailable. Left: schematic representation of the channel (in blue), membrane (yellow), and flow (arrows). Right: a snapshot of the M126V model during an MD trajectory, in cartoon representation in green. Internal space is represented by showing the solvent’s accessible surface in gray.
	Figure 3. Effects of the R351W mutation of SLC2A9b. Top: schematic representation of the channel (in blue), membrane (yellow), and flow (arrows). Bottom: snapshots of the R351W model during an MD trajectory in cartoon representation in green. Internal space is represented by showing the solvent accessible surface in gray (compared to similar surfaces describing the vestibular areas in Figure 2). The initial model structure is surprisingly stable, but it deforms under molecular dynamics simulations, as seen in the snapshot on the right after ~30 ns of MD trajectory. Notice the very substantial reorganization of the internal helices.
	Figure 4. Expression of WT and mutant SLC2A9b in Xenopus oocytes. (A) Schematic representation of the experimental procedure. The same amount of wild-type or mutants SLC2A9b RNAs were injected into oocytes. The oocytes were harvested after 2 days and then, Western blot analysis or immunostaining was performed. (B) SLC2A9b wild type and mutants showed similar protein expression levels. The histogram depicts the relative protein expression level (n = 3). Quantification with one-way ANOVA (Dunnett’s multiple comparisons test), p = 0.4363. Data represent the mean ± S.D. of three individual experiments. ns: no statistical differences between the groups. (C) Immunostaining was performed using anti-V5 antibodies. Both WT and mutants SLC2A9b showed plasma membrane localization in Xenopus oocytes. DIC; differential interference contrast image.
	Figure 5. Met126Val mutation reduces uric acid uptake. Uric acid uptake assay was performed as described in the Methods section. M126V mutation in SLC2A9b reduced uric acid uptake by 45% and R351W mutation decreased by 70%. Histogram depicts relative urate uptake level (n = 10). Quantification with one-way ANOVA (Dunnett’s multiple comparisons test), ** p < 0.0001. Data represent the mean ± S.D. of three individual experiments. ** p < 0.0001.

References [+] :

Adzhubei, A method and server for predicting damaging missense mutations. 2010, Pubmed