Identification and Characterization of Single Nucleotide Polymorphisms of SLC22A11 (hOAT4) in Korean Women Osteoporosis Patients

  • Lee, Woon Kyu (Department of Laboratory Animal Medicine, Medical Research Center, College of Medicine, Yonsei University) ;
  • Kwak, Jin Oh (Department of Pharmacology and Toxicology, Inha University) ;
  • Hwang, Ji-Sun (Department of Physiology and Biophysics and Center for Advanced Medical Education by BK21 Project, College of Medicine, Inha University) ;
  • Suh, Chang Kook (Department of Physiology and Biophysics and Center for Advanced Medical Education by BK21 Project, College of Medicine, Inha University) ;
  • Cha, Seok Ho (Department of Pharmacology and Toxicology, Inha University)
  • Received : 2007.10.16
  • Accepted : 2007.10.25
  • Published : 2008.04.30

Abstract

Single nucleotide polymorphisms (SNPs) are the most common form of human genetic variation. Non-synonymous SNPs (nsSNPs) change an amino acid. Organic anion transporters (OATs) play an important role in eliminating or reabsorbing endogenous and exogenous organic anionic compounds. Among OATs, hOAT4 mediates high affinity transport of estrone sulfate and dehydroepiandrosterone sulfate. The rapid bone loss that occurs in post-menopausal women is mainly due to a net decrease of estrogen. In the present study we searched for SNPs within the exon regions of hOAT4 in Korean women osteoporosis patients. Fifty healthy subjects and 50 subjects with osteoporosis were screened for genetic polymorphism in the coding region of SLC22A11 (hOAT4) using GC-clamp PCR and denaturing gradient gel electrophoresis (DGGE). We found three SNPs in the hOAT4 gene. Two were in the osteoporosis group (C483A and G832A) and one in the normal group (C847T). One of the SNPs, G832A, is an nsSNP that changes the $278^{th}$ amino acid from glutamic acid to lysine (E278K). Uptake of [$3^H$] estrone sulfate by oocytes injected with the hOAT4 E278K mutant was reduced compared with wild-type hOAT4. Km values for wild type and E278K were $0.7{\mu}M$ and $1.2{\mu}M$, and Vmax values were 1.8 and 0.47 pmol/oocyte/h, respectively. The present study demonstrates that hOAT4 variants can causing inter-individual variation in anionic drug uptake and, therefore, could be used as markers for certain diseases including osteoporosis.

Keywords

Denaturing Gradient Gel Electrophoresis;Estrone Sulfate;GC-Clamp;Human Organic Anion Transporter 4;Osteoporosis;Polymorphism

Acknowledgement

Supported by : Korea Research Foundation

References

  1. Babu, E., Takeda, M., Narikawa, S., Kobayashi, Y., Enomoto, A., Tojo, A., Cha, S.H., Sekine, T., Sakthisekaran, D., and Endou, H. (2002). Role of human organic anion transporter 4 in transport of ochratoxin A. Biochim. Biophys. Acta 1590, 64-75 https://doi.org/10.1016/S0167-4889(02)00187-8
  2. Bacsi, K., Kosa, J., Horvath, H., Balla, B., Lakatos, P., and Speer, G. (2007). Significance of dehydroepiandrosterone and dehydroepiandrosterone sulfate in different diseases. Orv. Hetil. 148, 651-657 https://doi.org/10.1556/OH.2007.27903
  3. Keitel, V., Nies, A.T., Brom, M., Hummel-Eisenbeiss, J., Spring, H., and Keppler, D. (2003). A common Dubin-Johnson syndrome mutation impairs protein maturation and transport activity of MRP2 (ABCC2). Am. J. Physiol. Gastrointest. Liver Physiol. 284, G165-G174 https://doi.org/10.1152/ajpgi.00362.2002
  4. Mumm, S., Jones, J., Finnegan, P., Menthorn, P.S., Podgomik, M.N., and Whyte, M.P. (2002). Denaturing gradient gel electrophoresis analysis of the tissue nonspecific alkaline phosphatase isoenzyme gene in hypophosphatasia. Mol. Genet. Metab. 75, 143-153 https://doi.org/10.1006/mgme.2001.3283
  5. Nozawa, T., Minami, H., Sugiura, S., Tsuji A., and Tamai, I. (2005). Role of organic anion transporter OATP1B1 (OATPC) in hepatic uptake of irinotecan and its active metabolite, 7-ethyl-10-hydroxycamptothecin: in vitro evidence and effect of single nucleotide polymorphisms. Drug Metab. Dispos. 33, 434-439 https://doi.org/10.1124/dmd.104.001909
  6. Trauner, M., and Boyer, J.L. (2003). Bile salt transporters: molecular characterization, function, and regulation. Physiol. Rev. 83, 633-671 https://doi.org/10.1152/physrev.00027.2002
  7. Wu, Y., Hayes, V.M., Osinga, J., Mulder, I.M., Looman, M.W., Buys, C.H., and Hofstra, R.M. (1998). Improvement of fragment and primer selection for mutation detection by denaturing gradient gel electrophoresis. Nucleic Acids Res. 26, 5432-5440 https://doi.org/10.1093/nar/26.23.5432
  8. Lee, S.S., Jeong, H.E., Yi, J.M., Jung, H.J., Jang, J.E., Kim, E.Y., Lee, S.J., and Shin, J.G. (2007). Identification and functional assessment of BCRP polymorphisms in a Korean population. Drug Metab. Dispos. 35, 623-632 https://doi.org/10.1124/dmd.106.012302
  9. Patel, P., Weerasekera, N., Hitchins, M., Boyd, C.A., Johnston, D.G., and Williamson, C. (2003). Semi quantitative expression analysis of MDR3, FIC1, BSEP, OATP-A, OATP-C, OATP-D, OATP-E and NTCP gene transcripts in 1st and 3rd trimester human placenta. Placenta 24, 39-44 https://doi.org/10.1053/plac.2002.0879
  10. Xu, G., Bhatnagar, V., Wen, G., Hamilton, B.A., Eraly, S.A., and Nigam, S.K. (2005). Analyses of coding region polymorphisms in apical and basolateral human organic anion transporter (OAT) genes OAT1 (NKT), OAT2, OAT3, OAT4, URAT (RST) . Kidney Int. 68, 1491-1499 https://doi.org/10.1111/j.1523-1755.2005.00612.x
  11. Van Orsouw, N.J., and Vijg, J. (1999). Design and application of 2-D DGGE-based gene mutational scanning test. Genet. Anal. 14, 205-213 https://doi.org/10.1016/S1050-3862(98)00028-X
  12. Riggs, B.L., Khosla, S., and Melton, L.J.3rd. (2002). Sex steroids and the construction and conservation of the adult skeleton. Endocr. Rev. 23, 279-302 https://doi.org/10.1210/er.23.3.279
  13. Wang, F.S., Ko, J.Y., Lin, C.L., Wu, H.L., Ke, H.J., and Tai, P.J. (2007). Knocking down dickkopf-1 alleviates estrogen deficiency induction of bone loss. A histomorphological study in ovariectomized rats. Bone 40, 485-492 https://doi.org/10.1016/j.bone.2006.09.004
  14. Briz, O., Serrano, M.A., MacIas, R.I., Gonzalez-Gallego, J., and Marin, J.J. (2003). Role of organic anion-transporting polypeptides, OATP-A, OATP-C and OATP-8, in the human placenta- maternal liver tandem excretory pathway for foetal bilirubin. Biochem. J. 371, 897-905 https://doi.org/10.1042/BJ20030034
  15. Evseenko, D.A., Murthi, P., Paxton, J.W., Reid, G., Emerald, B.S., Mohankumar, K.M., Lobie, P.E., Brennecke, S.P., and Keelan, J.A. (2007a). The ABC transporter BCRP/ABCG2 is a placental survival factor, and its expression is reduced in indiopathic human fetal growth restriction. FASEB J. 21, 3592-3605 https://doi.org/10.1096/fj.07-8688com
  16. Deighton, C.M., Walker, D.J., Griffiths, I.D., and Roberts, D.F. (1989). The contribution of HLA to rheumatoid arthritis. Clin. Genet. 36, 178-182 https://doi.org/10.1111/j.1399-0004.1989.tb03185.x
  17. Boyle, W.J., Simonet, W.S., and Lacey, D.L. (2003). Osteoclast differentiation and activation. Nature 423, 337-342 https://doi.org/10.1038/nature01658
  18. Evseenko, D.A., Paxton, J.W., and Keelan, J.A. (2007b). Independent regulation of apical and basolateral drug transporter expression and function in placental trophoblasts by cytokines, steroids, and growth factor. Drug Metab. Dispos. 35, 595-601 https://doi.org/10.1124/dmd.106.011478
  19. Jamroziak, K., Balcerczak, E., Smolewski, P., Robey, R.W., Cebula, B., Panczyk, M., Kowalczyk, M., Szmigielska- Kapłon, A., Mirowski, M., Bates, S.E., et al. (2006). MDR1 (ABCB1) gene polymorphism C3435T is associated with Pglycoprotein activity in B-cell chronic lymphocytic leukemia. Pharmacol. Rep. 58, 720-728
  20. Masi, A.T., Aldag, J.C., and Chatterton, R.T. (2006). Sex hormones and risks of rheumatoid arthritis and developmental or environmental influences. Ann. NY Acad. Sci. 1069, 223-235 https://doi.org/10.1196/annals.1351.020
  21. Firestein, G.S. (2003). Evolving concepts of rheumatoid arthritis. Nature 423, 356-361 https://doi.org/10.1038/nature01661
  22. Ho, R.H., Leake, B.F., Roberts, R.L., Lee, W., and Kim, R.B. (2004). Ethnicity-dependent polymorphism in$Na^+$-taurocholate cotransporting polypeptide (SLC10A1) reveals a domain critical for bile acid substrate recognition. J. Biol. Chem. 20, 7213-7222
  23. Bonjour, J.P., Theintz, G., Law, F., Slosman, D., and Rizzoli, R. (1995). Peak bone mass: facts and uncertainties. Arch. Pediatr. 2, 460-468 https://doi.org/10.1016/0929-693X(96)81183-3
  24. Evseenko, D.A., Paxton, J.W., and Keelan, J.A. (2006). ABC drug transporter expression and functional activity in trophoblast- like cell line and differentiating primary trophoblast. Am. J. Physiol. Regul. Integr. Comp. Physiol. 290, R1357-R1365 https://doi.org/10.1152/ajpregu.00630.2005
  25. Cha, S.H., Sekine, T., Fukushima, J.I., Kanai, Y., Kobayashi, Y., Goya, T., and Endou, H. (2001). Identification and characterization of human organic anion transporter 3 expressing predominantly in the kidney. Mol. Pharm. 59, 1277-1286 https://doi.org/10.1124/mol.59.5.1277
  26. Ugele, B., St-Pierre, M.V., Pihusch, M., Bahn, A., and Hantschmann, P. (2003). Characterization and identification of steroid sulfate transporters of human placenta. Am. J. Physiol. Endocrinol. Metab. 284, E390-E398 https://doi.org/10.1152/ajpendo.00257.2002
  27. Cha, S.H., Sekine, T., Kusuhara, H., Yu, E., Kim, J.Y., Kim, D.K., Sugiyama, Y., Kanai, Y., and Endou, H. (2000). Molecular cloning and characterization of multispecific organic anion transporter 4 expressed in the placenta. J. Biol. Chem. 275, 4507-4512 https://doi.org/10.1074/jbc.275.6.4507
  28. Klareskog, L., Padyukov, L., Lorentzen, J., and Alfredsson, L. (2006). Mechanisms of disease: genetic susceptibility and environmental triggers in the development of rheumatoid arthritis. Nat. Clin. Pract. Rheumatol. 2, 425-433 https://doi.org/10.1038/ncprheum0249
  29. Conrad, S., Kauffmann, H.M., Ito, K., Deeley, R.G., Cole, S.P., and Schrenk, D. (2001). Identification of human multidrug resistance protein 1 (MRP1) mutations and characterization of a G671V substitution. J. Hum. Genet. 46, 656-663 https://doi.org/10.1007/s100380170017
  30. Lee, J.H., Jeong, S.M., Kim, J.H., Lee, B.H., Yoon, I.S., Lee, J.H., Choi, S.H., Lee, S.M., Park, Y.S., Lee, J.H., et al. (2006). Effects of ginsenosides and their metabolites on voltage- dependent $Ca^2+$ channel sybtypes. Mol. Cells 21, 52-62
  31. Wang, H., Wu, X., Hudkins, K., Mikheev, A., Zhang, H., Gupta, A., Unadkat, J.D., and Mao, Q. (2006). Expression of the breast cancer resistance protein (Bcrp1/Abcg2) in tissues from preg- nant mice: effects of pregnancy and correlations with nuclear receptors. Am. J. Physiol. Endocrinol. Metab. 291, E1295-E1303 https://doi.org/10.1152/ajpendo.00193.2006