title: In Vitro Transport Activity and Trafficking of MRP2/ABCC2 Polymorphic Variants.
abstract: OBJECTIVE
  Multidrug resistance-associated protein 2 (MRP2/ABCC2) is an efflux pump that removes drugs and chemicals from cells. We sought to characterize the expression, trafficking and in vitro activity of seven single nucleotide polymorphisms (SNPs) in the ABCC2 gene.
  
  
  METHODS
  ABCC2 SNPs were generated using site-directed mutagenesis and stably expressed in Flp-In HEK293 cells, which allows targeted insertion of transgenes within the genome. Total and cell surface expression of MRP2 as well as accumulation of substrates (calcein AM and 5(6)-carboxy-2',7'-dichlorofluorescein diacetate, CDCF) were quantified in cells or inverted membrane vesicles expressing wild-type (WT) or variant forms.
  
  
  RESULTS
  The cell surface expression of the C-24T-, G1249A-, G3542T-, T3563A-, C3972T- and G4544A-MRP2 variants was similar to WT-MRP2. While expression was similar, transport of calcein AM was enhanced in cells expressing the G3542T-, T3563A-, C3972T-, and G4544A-MRP2 variants. By comparison, cells expressing the C2366T-MRP2 variant had 40-50% lower surface expression, which increased the accumulation of calcein AM up to 3-fold. Accumulation of CDCF in inverted membrane vesicles expressing the C2366T-MRP2 variant was also reduced by 50%. In addition, the G1249A-MRP2 variant had decreased transport of CDCF.
  
  
  CONCLUSIONS
  Taken together, these data demonstrate that genetic variability in the ABCC2 gene influences the in vitro expression, trafficking, and transport activity of MRP2.
authors.affiliation: Department of Pharmacology and Toxicology, Rutgers University Ernest Mario School of Pharmacy, 170 Frelinghuysen Road, Piscataway, New Jersey, 08854, USA.
authors.forename: Xia
authors.initials: X
authors.lastname: Wen
authors.affiliation: Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado, Aurora, Colorado, 80045, USA.
authors.forename: Melanie S
authors.initials: MS
authors.lastname: Joy
authors.affiliation: Department of Pharmacology and Toxicology, Rutgers University Ernest Mario School of Pharmacy, 170 Frelinghuysen Road, Piscataway, New Jersey, 08854, USA. aleksunes@eohsi.rutgers.edu.|Environmental and Occupational Health Sciences Institute, Rutgers University, Piscataway, New Jersey, 08854, USA. aleksunes@eohsi.rutgers.edu.
authors.forename: Lauren M
authors.initials: LM
authors.lastname: Aleksunes
mesh_terms: D015870:Gene Expression; D014644:Genetic Variation; D057809:HEK293 Cells; D006801:Humans; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D020641:Polymorphism, Single Nucleotide; D021381:Protein Transport
chemical_list: D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; C000715087:ABCC2 protein, human
keywords: ABCC2; MRP2; SNP; transporter; variant