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Polymorphisms of SLC22A9 (hOAT7) in Korean Females with Osteoporosis.

Ahn SK, Suh CK, Cha SH - Korean J. Physiol. Pharmacol. (2015)

Bottom Line: The net decrease of estrogen, especially in post-menopausal women induces rapid bone loss.The G767A, T1277C and C1487T SNPs result in an amino acid substitution, from synonymous vs nonsynonymous substitution arginine to glutamine (R256Q), phenylalanine to serine (F426S) and proline to leucine (P496L), respectively.The Km values and Vmax of the wild type, R256Q, P496L and F426S were 8.84, 8.87, 9.83 and 12.74 µM, and 1.97, 1.96, 2.06 and 1.55 pmol/oocyte/h, respectively.

View Article: PubMed Central - PubMed

Affiliation: Department of Tropical Medicine and Parasitology, College of Medicine, Inha University, Incheon 400-712, Korea.

ABSTRACT
Among solute carrier proteins, the organic anion transporters (OATs) play an important role for the elimination or reabsorption of endogenous and exogenous negatively charged anionic compounds. Among OATs, SLC22A9 (hOAT7) transports estrone sulfate with high affinity. The net decrease of estrogen, especially in post-menopausal women induces rapid bone loss. The present study was performed to search the SNP within exon regions of SLC22A9 in Korean females with osteoporosis. Fifty healthy controls and 50 osteoporosis patients were screened for the genetic polymorphism in the coding region of SLC22A9 using GC-clamped PCR and denaturing gradient gel electrophoresis (DGGE). Six SNPs were found on the SLC22A9 gene from Korean women with/without osteoporosis. The SNPs were located as follows: two SNPs in the osteoporosis group (A645G and T1277C), three SNPs in the control group (G1449T, C1467T and C1487T) and one SNP in both the osteoporosis and control groups (G767A). The G767A, T1277C and C1487T SNPs result in an amino acid substitution, from synonymous vs nonsynonymous substitution arginine to glutamine (R256Q), phenylalanine to serine (F426S) and proline to leucine (P496L), respectively. The Km values and Vmax of the wild type, R256Q, P496L and F426S were 8.84, 8.87, 9.83 and 12.74 µM, and 1.97, 1.96, 2.06 and 1.55 pmol/oocyte/h, respectively. The present study demonstrates that the SLC22A9 variant F426S is causing inter-individual variation that is leading to the differences in transport of the steroid sulfate conjugate (estrone sulfate) and, therefore this could be used as a marker for certain disease including osteoporosis.

No MeSH data available.


Related in: MedlinePlus

Transport activity of the SLC22A9 and its variants. (A) Concentration dependence of estrone sulfate mediated by wild type SLC22A9 and variants in Xenopus laevis oocytes. Defolliculated stage VI and V oocytes were injected with 25 ng/oocyte of wild-type and variants cRNAs, and incubated for 3 days in Barth's solution at 18℃. After three days, [3H] estrone sulfate uptake experiment was performed using various concentrations (25 nM~50 µM). Each point employs 8~10 oocytes and represented mean±standard error of three independent experiments. (B) Eadie-Hofstee plot analysis for determination of Km and Vmax values.
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Figure 3: Transport activity of the SLC22A9 and its variants. (A) Concentration dependence of estrone sulfate mediated by wild type SLC22A9 and variants in Xenopus laevis oocytes. Defolliculated stage VI and V oocytes were injected with 25 ng/oocyte of wild-type and variants cRNAs, and incubated for 3 days in Barth's solution at 18℃. After three days, [3H] estrone sulfate uptake experiment was performed using various concentrations (25 nM~50 µM). Each point employs 8~10 oocytes and represented mean±standard error of three independent experiments. (B) Eadie-Hofstee plot analysis for determination of Km and Vmax values.

Mentions: In order to examine whether the SLC22A9 polymorphism found in Korean osteoporosis patients affects the functional activity on substrate transport, we constructed mutant cDNAs and expressed them into Xenopus oocytes. As shown in Fig. 3, F426S exhibited reduced uptake for [3H] estrone sulfate compared with that of the wild-type SLC22A9. R256Q and P496L did not show a statistically significant change in [3H] estrone sulfate compared with those of the wild-type SLC22A9. According to the changes of concentration-dependent [3H] estrone sulfate in wild type and F426S, the Km values for wild-type and F426S were 0.7 and 1.2 µM, respectively, and the Vmax values for wildtype and F426S were 1.8 and 0.47 pmol/oocyte/h, respectively (Fig. 3).


Polymorphisms of SLC22A9 (hOAT7) in Korean Females with Osteoporosis.

Ahn SK, Suh CK, Cha SH - Korean J. Physiol. Pharmacol. (2015)

Transport activity of the SLC22A9 and its variants. (A) Concentration dependence of estrone sulfate mediated by wild type SLC22A9 and variants in Xenopus laevis oocytes. Defolliculated stage VI and V oocytes were injected with 25 ng/oocyte of wild-type and variants cRNAs, and incubated for 3 days in Barth's solution at 18℃. After three days, [3H] estrone sulfate uptake experiment was performed using various concentrations (25 nM~50 µM). Each point employs 8~10 oocytes and represented mean±standard error of three independent experiments. (B) Eadie-Hofstee plot analysis for determination of Km and Vmax values.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4499643&req=5

Figure 3: Transport activity of the SLC22A9 and its variants. (A) Concentration dependence of estrone sulfate mediated by wild type SLC22A9 and variants in Xenopus laevis oocytes. Defolliculated stage VI and V oocytes were injected with 25 ng/oocyte of wild-type and variants cRNAs, and incubated for 3 days in Barth's solution at 18℃. After three days, [3H] estrone sulfate uptake experiment was performed using various concentrations (25 nM~50 µM). Each point employs 8~10 oocytes and represented mean±standard error of three independent experiments. (B) Eadie-Hofstee plot analysis for determination of Km and Vmax values.
Mentions: In order to examine whether the SLC22A9 polymorphism found in Korean osteoporosis patients affects the functional activity on substrate transport, we constructed mutant cDNAs and expressed them into Xenopus oocytes. As shown in Fig. 3, F426S exhibited reduced uptake for [3H] estrone sulfate compared with that of the wild-type SLC22A9. R256Q and P496L did not show a statistically significant change in [3H] estrone sulfate compared with those of the wild-type SLC22A9. According to the changes of concentration-dependent [3H] estrone sulfate in wild type and F426S, the Km values for wild-type and F426S were 0.7 and 1.2 µM, respectively, and the Vmax values for wildtype and F426S were 1.8 and 0.47 pmol/oocyte/h, respectively (Fig. 3).

Bottom Line: The net decrease of estrogen, especially in post-menopausal women induces rapid bone loss.The G767A, T1277C and C1487T SNPs result in an amino acid substitution, from synonymous vs nonsynonymous substitution arginine to glutamine (R256Q), phenylalanine to serine (F426S) and proline to leucine (P496L), respectively.The Km values and Vmax of the wild type, R256Q, P496L and F426S were 8.84, 8.87, 9.83 and 12.74 µM, and 1.97, 1.96, 2.06 and 1.55 pmol/oocyte/h, respectively.

View Article: PubMed Central - PubMed

Affiliation: Department of Tropical Medicine and Parasitology, College of Medicine, Inha University, Incheon 400-712, Korea.

ABSTRACT
Among solute carrier proteins, the organic anion transporters (OATs) play an important role for the elimination or reabsorption of endogenous and exogenous negatively charged anionic compounds. Among OATs, SLC22A9 (hOAT7) transports estrone sulfate with high affinity. The net decrease of estrogen, especially in post-menopausal women induces rapid bone loss. The present study was performed to search the SNP within exon regions of SLC22A9 in Korean females with osteoporosis. Fifty healthy controls and 50 osteoporosis patients were screened for the genetic polymorphism in the coding region of SLC22A9 using GC-clamped PCR and denaturing gradient gel electrophoresis (DGGE). Six SNPs were found on the SLC22A9 gene from Korean women with/without osteoporosis. The SNPs were located as follows: two SNPs in the osteoporosis group (A645G and T1277C), three SNPs in the control group (G1449T, C1467T and C1487T) and one SNP in both the osteoporosis and control groups (G767A). The G767A, T1277C and C1487T SNPs result in an amino acid substitution, from synonymous vs nonsynonymous substitution arginine to glutamine (R256Q), phenylalanine to serine (F426S) and proline to leucine (P496L), respectively. The Km values and Vmax of the wild type, R256Q, P496L and F426S were 8.84, 8.87, 9.83 and 12.74 µM, and 1.97, 1.96, 2.06 and 1.55 pmol/oocyte/h, respectively. The present study demonstrates that the SLC22A9 variant F426S is causing inter-individual variation that is leading to the differences in transport of the steroid sulfate conjugate (estrone sulfate) and, therefore this could be used as a marker for certain disease including osteoporosis.

No MeSH data available.


Related in: MedlinePlus