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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

Efflux of [125I]T3 and [125I]T4 by WT and mutated MCT8.Efflux of [125I]T3 (A,C,E) and [125I]T4 (B,D,F) for 2–30 min from fibroblasts (A,B), and transiently transfected COS1 cells (C,D) and JEG3 cells (E,F). Efflux is shown as percentage of cellular radioactivity at 0 min and is corrected for protein. Results are presented as mean ± SEM (n = 3). Significance is indicated for control vs. patient fibroblast (A,B) or mutual difference between WT MCT8 and mutants. *P <0.05; **P <0.01; ***P <0.001.
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pone.0139343.g005: Efflux of [125I]T3 and [125I]T4 by WT and mutated MCT8.Efflux of [125I]T3 (A,C,E) and [125I]T4 (B,D,F) for 2–30 min from fibroblasts (A,B), and transiently transfected COS1 cells (C,D) and JEG3 cells (E,F). Efflux is shown as percentage of cellular radioactivity at 0 min and is corrected for protein. Results are presented as mean ± SEM (n = 3). Significance is indicated for control vs. patient fibroblast (A,B) or mutual difference between WT MCT8 and mutants. *P <0.05; **P <0.01; ***P <0.001.

Mentions: Because the fibroblast uptake studies suggested impaired T3 and T4 efflux from the patient’s cells, we decided to study [125I]T3 and [125I]T4 efflux directly from patient’s and control fibroblasts. Fig 5A shows a significantly impaired T3 efflux from patient’s fibroblasts. T4 efflux was also reduced but not to the same degree as T3 efflux (Fig 5B). Only at 10 minutes a significant difference was found with T4 efflux from control fibroblasts. In contrast, T3 and T4 efflux were not significantly different between COS1 or JEG3 cells transiently transfected with WT MCT or the S290F mutant (Fig 5C–5F). The S290A mutant was not significantly different from WT MCT8 in mediating T3 and T4 efflux from both cell types.


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)

Efflux of [125I]T3 and [125I]T4 by WT and mutated MCT8.Efflux of [125I]T3 (A,C,E) and [125I]T4 (B,D,F) for 2–30 min from fibroblasts (A,B), and transiently transfected COS1 cells (C,D) and JEG3 cells (E,F). Efflux is shown as percentage of cellular radioactivity at 0 min and is corrected for protein. Results are presented as mean ± SEM (n = 3). Significance is indicated for control vs. patient fibroblast (A,B) or mutual difference between WT MCT8 and mutants. *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.g005: Efflux of [125I]T3 and [125I]T4 by WT and mutated MCT8.Efflux of [125I]T3 (A,C,E) and [125I]T4 (B,D,F) for 2–30 min from fibroblasts (A,B), and transiently transfected COS1 cells (C,D) and JEG3 cells (E,F). Efflux is shown as percentage of cellular radioactivity at 0 min and is corrected for protein. Results are presented as mean ± SEM (n = 3). Significance is indicated for control vs. patient fibroblast (A,B) or mutual difference between WT MCT8 and mutants. *P <0.05; **P <0.01; ***P <0.001.
Mentions: Because the fibroblast uptake studies suggested impaired T3 and T4 efflux from the patient’s cells, we decided to study [125I]T3 and [125I]T4 efflux directly from patient’s and control fibroblasts. Fig 5A shows a significantly impaired T3 efflux from patient’s fibroblasts. T4 efflux was also reduced but not to the same degree as T3 efflux (Fig 5B). Only at 10 minutes a significant difference was found with T4 efflux from control fibroblasts. In contrast, T3 and T4 efflux were not significantly different between COS1 or JEG3 cells transiently transfected with WT MCT or the S290F mutant (Fig 5C–5F). The S290A mutant was not significantly different from WT MCT8 in mediating T3 and T4 efflux from both cell types.

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