Genetic polymorphisms in the multidrug resistance-associated protein 3 (ABCC3, MRP3) gene and relationship to its mRNA and protein expression in human liver.
OBJECTIVE
To determine the genetic variability of multidrug resistance protein 3 (MRP3).


METHODS
Genomic DNA samples from 103 Caucasians were systematically screened for genetic variations to find a potential relationship with hepatic MRP3 expression. Sequencing comprised all 31 exons, approximately 100 bp of the flanking intronic regions and 2 kb of the 5' UTR.


RESULTS
In total, 51 mutations were identified. Fifteen SNPs were located in the coding exons of MRP3, six of which are nonsynonymous mutations. SNPs 39G>C (allele frequency: 0.5%, located in exon 1), 202C>T (1.6%, exon 2), 1037C>T (0.5%, exon 9), 1537C>A (0.5%, exon 12), 3890G>A (5.2%, exon 27) and 4267G>A (0.6%, exon 29) resulted in Lys13Asn, His68Tyr, Ser346Phe, Gln513Lys, Arg1297His and Gly1423Arg amino acid substitutions, respectively. A splice site mutation (1339-1G>T) was found at the intron 10-exon 11 boundary. To evaluate, whether mutations in the MRP3 gene correlate with human hepatic MRP3 expression, we analyzed the genetic variants in Caucasian liver samples, whose MRP3 mRNA (n = 84) and protein (n = 50) expression has been determined by real time quantitative PCR and Western Blot, respectively. We found a significant correlation of a polymorphism in the 5' promoter region (-211C>T) of MRP3 with mRNA expression. Individuals homozygous and heterozygous for the -211C>T promoter polymorphism had significantly lower MRP3 transcript levels compared to wild-type individuals (P < 0.05). Accordingly, electrophoretic mobility shift assay demonstrated that -211C>T polymorphism affected the binding of nuclear factors.


CONCLUSIONS
Multiple genetic polymorphisms of MRP3 exist in Caucasians. The -211C>T promoter polymorphism appears to be associated with altered hepatic MRP3 mRNA expression.
Lang
Thomas
T
Epidauros Biotechnology, Pharmacogenetics Laboratory, Am Neuland 1, 82347 Bernried, Germany.
Hitzl
Monika
M
Burk
Oliver
O
Mornhinweg
Esther
E
Keil
Andrea
A
Kerb
Reinhold
R
Klein
Kathrin
K
Zanger
Ulrich M
UM
Eichelbaum
Michel
M
Fromm
Martin F
MF
D017398:Alternative Splicing; D019943:Amino Acid Substitution; D015153:Blotting, Western; D018432:Drug Resistance, Multiple; D024202:Electrophoretic Mobility Shift Assay; D005091:Exons; D005787:Gene Frequency; D006579:Heterozygote; D006720:Homozygote; D006801:Humans; D008099:Liver / Q000378:metabolism / Q000473:pathology; D008113:Liver Neoplasms / Q000235:genetics* / Q000378:metabolism* / Q000556:secondary; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000235:genetics* / Q000378:metabolism*; D009154:Mutation; D009687:Nuclear Proteins / Q000378:metabolism; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide; D011401:Promoter Regions, Genetic; D044465:White People
D027425:ATP-Binding Cassette, Sub-Family C Proteins; D009687:Nuclear Proteins; C114462:multidrug resistance-associated protein 3