title: Functional characterization of nonsynonymous single nucleotide polymorphisms in the human organic anion transporter 4 (hOAT4).
abstract: OBJECTIVE
  The human organic anion transporter (hOAT) family of transmembrane carrier proteins mediate the cellular flux of anionic substances, including certain hormones and anti-cancer drugs. hOAT4 is highly expressed at the apical membrane of the renal tubular cell and facilitates drug re-absorption in the kidney. In the present study, the impact of 10 nonsynonymous single nucleotide polymorphisms (SNPs) of hOAT4 on transport function in COS-7 cells was characterized.
  
  
  METHODS
  Transport uptake assay was used to assess the function of the variant transporters. Cell surface biotinylation and western blot analysis were used to investigate the expression characteristics of the transporter proteins. Comparative modelling was used to interpret the influence of nonsynonymous changes in terms of hOAT4 structure.
  
  
  RESULTS
  Four naturally occurring hOAT4 variants (L29P, R48Y, V155G and T392I) exhibited a significant loss of function. Substitution of leucine-29, which is a conserved residue in OATs, with a proline residue, impaired the synthesis or the apparent stability of the transporter and membrane insertion was disrupted in the R48Y variant. In the case of the V155G and T392I variants, impaired function was due to decreased affinity of the transporter for oestrone sulphate and impaired transporter-substrate turnover respectively. The T392I variant was inhibited more extensively than the wild-type transporter by the cationic substrate tetraethyl ammonium.
  
  
  CONCLUSIONS
  Several naturally occurring SNPs encode variant hOAT4s that may impair the renal tubular re-absorption of important drug substrates.
authors.affiliation: Pharmacogenomics and Drug Development Laboratory, Faculty of Pharmacy, The University of Sydney, Sydney, NSW, Australia. fzhou@pharm.usyd.edu.au
authors.forename: Fanfan
authors.initials: F
authors.lastname: Zhou
authors.forename: Ling
authors.initials: L
authors.lastname: Zhu
authors.forename: Pei H
authors.initials: PH
authors.lastname: Cui
authors.forename: W Bret
authors.initials: WB
authors.lastname: Church
authors.forename: Michael
authors.initials: M
authors.lastname: Murray
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D019556:COS Cells; D002462:Cell Membrane / Q000378:metabolism; D002522:Chlorocebus aethiops; D004970:Estrone / Q000031:analogs & derivatives / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D029362:Organic Anion Transporters, Sodium-Independent / Q000235:genetics* / Q000378:metabolism; D020641:Polymorphism, Single Nucleotide
chemical_list: D029362:Organic Anion Transporters, Sodium-Independent; C427207:SLC22A11 protein, human; D004970:Estrone; C017296:estrone sulfate