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Further Insights into the Allan-Herndon-Dudley Syndrome: Clinical and Functional Characterization of a Novel MCT8 Mutation.

Armour CM, Kersseboom S, Yoon G, Visser TJ - PLoS ONE (2015)

Bottom Line: Functional analysis of the S290F mutant showed decreased TH transport, metabolism and protein expression in the three cell types, whereas the S290A mutation had no effect.However, no effect of the S290F mutation was observed on TH efflux from COS1 and JEG3 cells.Furthermore, our results indicate that the function of the S290F mutant is dependent on cell context.

View Article: PubMed Central - PubMed

Affiliation: Regional Genetics Program, Children's Hospital of Eastern Ontario, and Children's Hospital of Eastern Ontario Research Institute, University of Ottawa, Ottawa, Canada.

ABSTRACT

Background: Mutations in the thyroid hormone (TH) transporter MCT8 have been identified as the cause for Allan-Herndon-Dudley Syndrome (AHDS), characterized by severe psychomotor retardation and altered TH serum levels. Here we report a novel MCT8 mutation identified in 4 generations of one family, and its functional characterization.

Methods: Proband and family members were screened for 60 genes involved in X-linked cognitive impairment and the MCT8 mutation was confirmed. Functional consequences of MCT8 mutations were studied by analysis of [125I]TH transport in fibroblasts and transiently transfected JEG3 and COS1 cells, and by subcellular localization of the transporter.

Results: The proband and a male cousin demonstrated clinical findings characteristic of AHDS. Serum analysis showed high T3, low rT3, and normal T4 and TSH levels in the proband. A MCT8 mutation (c.869C>T; p.S290F) was identified in the proband, his cousin, and several female carriers. Functional analysis of the S290F mutant showed decreased TH transport, metabolism and protein expression in the three cell types, whereas the S290A mutation had no effect. Interestingly, both uptake and efflux of T3 and T4 was impaired in fibroblasts of the proband, compared to his healthy brother. However, no effect of the S290F mutation was observed on TH efflux from COS1 and JEG3 cells. Immunocytochemistry showed plasma membrane localization of wild-type MCT8 and the S290A and S290F mutants in JEG3 cells.

Conclusions: We describe a novel MCT8 mutation (S290F) in 4 generations of a family with Allan-Herndon-Dudley Syndrome. Functional analysis demonstrates loss-of-function of the MCT8 transporter. Furthermore, our results indicate that the function of the S290F mutant is dependent on cell context. Comparison of the S290F and S290A mutants indicates that it is not the loss of Ser but its substitution with Phe, which leads to S290F dysfunction.

No MeSH data available.


Related in: MedlinePlus

Metabolism of [125I]T3 and [125I]T4 by D3 and T3 uptake saturation in MCT8 expressing cells.Metabolism of [125I]T3 (A) and [125I]T4 (B) after 4 hours in COS1 and JEG3 cells co-transfected with pCIneo-hD3 and empty vector, WT or mutant MCT8. Metabolism is shown as % of radioactivity in the incubation medium. C Inhibition of [125I]T3 uptake by WT and mutated MCT8 by increasing concentrations of unlabelled T3. Results are presented as mean ± SEM (n = 3). Significance is indicated for D3 co-transfected with WT or mutant MCT8 vs. D3 alone. *P <0.05; **P <0.01; ***P <0.001.
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pone.0139343.g006: Metabolism of [125I]T3 and [125I]T4 by D3 and T3 uptake saturation in MCT8 expressing cells.Metabolism of [125I]T3 (A) and [125I]T4 (B) after 4 hours in COS1 and JEG3 cells co-transfected with pCIneo-hD3 and empty vector, WT or mutant MCT8. Metabolism is shown as % of radioactivity in the incubation medium. C Inhibition of [125I]T3 uptake by WT and mutated MCT8 by increasing concentrations of unlabelled T3. Results are presented as mean ± SEM (n = 3). Significance is indicated for D3 co-transfected with WT or mutant MCT8 vs. D3 alone. *P <0.05; **P <0.01; ***P <0.001.

Mentions: To study the effects of the S290F and S290A mutations on the intracellular TH availability, we studied the metabolism of [125I]T3 and [125I]T4 in JEG3 and COS1 cells co-transfected with D3 and WT or mutant MCT8. Stimulation of intracellular deiodination by WT MCT8 was 2.9-fold for T3 and 8.1-fold for T4 in COS1 cells, and was 2.6-fold for T3 and 10.1-fold for T4 in JEG3 cells (Fig 6A and 6B). Relative to WT MCT8, the activity of the S290F mutant in facilitating intracellular deiodination was 85% for T3 and 74% for T4 in COS1 cells, and 47% for T3 and 35% for T4 in JEG3 cells. These findings indicate that the S290F mutant has relatively high activity in stimulating intracellular TH availability in COS1 cells, although low but significant facilitation of T3 and in particular T4 deiodination was also observed in JEG3 cells. No difference in facilitating intracellular TH metabolism was observed between WT MCT8 and the S290A mutant.


Further Insights into the Allan-Herndon-Dudley Syndrome: Clinical and Functional Characterization of a Novel MCT8 Mutation.

Armour CM, Kersseboom S, Yoon G, Visser TJ - PLoS ONE (2015)

Metabolism of [125I]T3 and [125I]T4 by D3 and T3 uptake saturation in MCT8 expressing cells.Metabolism of [125I]T3 (A) and [125I]T4 (B) after 4 hours in COS1 and JEG3 cells co-transfected with pCIneo-hD3 and empty vector, WT or mutant MCT8. Metabolism is shown as % of radioactivity in the incubation medium. C Inhibition of [125I]T3 uptake by WT and mutated MCT8 by increasing concentrations of unlabelled T3. Results are presented as mean ± SEM (n = 3). Significance is indicated for D3 co-transfected with WT or mutant MCT8 vs. D3 alone. *P <0.05; **P <0.01; ***P <0.001.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4591285&req=5

pone.0139343.g006: Metabolism of [125I]T3 and [125I]T4 by D3 and T3 uptake saturation in MCT8 expressing cells.Metabolism of [125I]T3 (A) and [125I]T4 (B) after 4 hours in COS1 and JEG3 cells co-transfected with pCIneo-hD3 and empty vector, WT or mutant MCT8. Metabolism is shown as % of radioactivity in the incubation medium. C Inhibition of [125I]T3 uptake by WT and mutated MCT8 by increasing concentrations of unlabelled T3. Results are presented as mean ± SEM (n = 3). Significance is indicated for D3 co-transfected with WT or mutant MCT8 vs. D3 alone. *P <0.05; **P <0.01; ***P <0.001.
Mentions: To study the effects of the S290F and S290A mutations on the intracellular TH availability, we studied the metabolism of [125I]T3 and [125I]T4 in JEG3 and COS1 cells co-transfected with D3 and WT or mutant MCT8. Stimulation of intracellular deiodination by WT MCT8 was 2.9-fold for T3 and 8.1-fold for T4 in COS1 cells, and was 2.6-fold for T3 and 10.1-fold for T4 in JEG3 cells (Fig 6A and 6B). Relative to WT MCT8, the activity of the S290F mutant in facilitating intracellular deiodination was 85% for T3 and 74% for T4 in COS1 cells, and 47% for T3 and 35% for T4 in JEG3 cells. These findings indicate that the S290F mutant has relatively high activity in stimulating intracellular TH availability in COS1 cells, although low but significant facilitation of T3 and in particular T4 deiodination was also observed in JEG3 cells. No difference in facilitating intracellular TH metabolism was observed between WT MCT8 and the S290A mutant.

Bottom Line: Functional analysis of the S290F mutant showed decreased TH transport, metabolism and protein expression in the three cell types, whereas the S290A mutation had no effect.However, no effect of the S290F mutation was observed on TH efflux from COS1 and JEG3 cells.Furthermore, our results indicate that the function of the S290F mutant is dependent on cell context.

View Article: PubMed Central - PubMed

Affiliation: Regional Genetics Program, Children's Hospital of Eastern Ontario, and Children's Hospital of Eastern Ontario Research Institute, University of Ottawa, Ottawa, Canada.

ABSTRACT

Background: Mutations in the thyroid hormone (TH) transporter MCT8 have been identified as the cause for Allan-Herndon-Dudley Syndrome (AHDS), characterized by severe psychomotor retardation and altered TH serum levels. Here we report a novel MCT8 mutation identified in 4 generations of one family, and its functional characterization.

Methods: Proband and family members were screened for 60 genes involved in X-linked cognitive impairment and the MCT8 mutation was confirmed. Functional consequences of MCT8 mutations were studied by analysis of [125I]TH transport in fibroblasts and transiently transfected JEG3 and COS1 cells, and by subcellular localization of the transporter.

Results: The proband and a male cousin demonstrated clinical findings characteristic of AHDS. Serum analysis showed high T3, low rT3, and normal T4 and TSH levels in the proband. A MCT8 mutation (c.869C>T; p.S290F) was identified in the proband, his cousin, and several female carriers. Functional analysis of the S290F mutant showed decreased TH transport, metabolism and protein expression in the three cell types, whereas the S290A mutation had no effect. Interestingly, both uptake and efflux of T3 and T4 was impaired in fibroblasts of the proband, compared to his healthy brother. However, no effect of the S290F mutation was observed on TH efflux from COS1 and JEG3 cells. Immunocytochemistry showed plasma membrane localization of wild-type MCT8 and the S290A and S290F mutants in JEG3 cells.

Conclusions: We describe a novel MCT8 mutation (S290F) in 4 generations of a family with Allan-Herndon-Dudley Syndrome. Functional analysis demonstrates loss-of-function of the MCT8 transporter. Furthermore, our results indicate that the function of the S290F mutant is dependent on cell context. Comparison of the S290F and S290A mutants indicates that it is not the loss of Ser but its substitution with Phe, which leads to S290F dysfunction.

No MeSH data available.


Related in: MedlinePlus