Relevance of Different Cellular Models in Determining the Effects of Mutations on SLC16A2/MCT8 Thyroid Hormone Transporter Function and Genotype-Phenotype Correlation

Abstract

SLC 16A2, the gene for the second member of the solute carrier family 16 (monocarboxylic acid transporter), located on chromosome Xq13.2, encodes a very efficient thyroid hormone transporter: monocarboxylate transporter 8, MCT8. Its loss of function is responsible in males for a continuum of psychomotor retardation ranging from severe (no motor acquisition, no speech) to mild (ability to walk with help and a few words of speech). Triiodothyronine uptake measurement in transfected cells and, more recently, patient fibroblasts, has been described to study the functional consequences of MCT8 mutations. Here, we describe three novel MCT8 mutations, including one missense variation not clearly predicted to be damaging but found in a severely affected patient. Functional studies in fibroblasts and JEG3 cells demonstrate the usefulness of both cellular models in validating the deleterious effects of a new MCT8 mutation if there is still a doubt as to its pathogenicity. Moreover, the screening of fibroblasts from a large number of patient fibroblasts and of transfected mutations has allowed us to demonstrate that JEG3 transfected cells are more relevant than fibroblasts in revealing a genotype-phenotype correlation.

Modulation of monocarboxylate transporter 8 oligomerization by specific pathogenic mutations

“…This therefore plays a decisive role in why a precise established relationship does not exist between the genotype, molecular characteristics of MCT8 variants (localization, transport routes), and the severity of patient phenotypes (Visser et al 2011) (summarized in Supplementary Table 1). The functional characteristics of the MCT8 mutants investigated in this current study were previously explored by in vitro analyses using JEG, MDCK, or COS cells (Schwartz et al 2005, Kinne et al 2009, 2010, Capri et al 2013, Groeneweg et al 2014. The cellular localization of MCT8 mutants and the rates of thyroid hormone uptake for all of these mutants finally revealed a diminished rate of T3 uptake.…”

“…Extensive functional in vitro characterization of MCT8 mutants revealed that the results obtained might vary depending on the cellular system used (Jansen et al 2005, Kinne et al 2009, Capri et al 2013. This therefore plays a decisive role in why a precise established relationship does not exist between the genotype, molecular characteristics of MCT8 variants (localization, transport routes), and the severity of patient phenotypes (Visser et al 2011) (summarized in Supplementary Table 1).…”

“…It is assumed that during transport of substrate, this 'bottle-neck' of the closed channel is opened by helical re-arrangements. A few of these WT amino acids are known to be involved in both substrate transport and intramolecular interaction, such as Arg445 (Kinne et al 2010, Capri et al 2013. Therefore, the tri-functionality of particular WT amino acids is important for: i) binding of substrate, ii) participating in intramolecular interaction network(s), and iii) switching the global protomer conformation.…”

Variable Clinical Characteristics and Molecular Spectrum of Patients with Syndromes of Reduced Sensitivity to Thyroid Hormone: Genetic Defects in the <b><i>THRB</i></b> and <b><i>SLC16A2</i></b> Genes

“…SIFT software predicted that SLC16A2 c.671C>T (p.A224V) and c.661G>A (p.G221R) would be tolerated. However, these variants were predicted to be deleterious by PolyPhen-2 and CADD score and proven to be pathogenic by in vitro functional analysis [19,20]. The p.W398R mutation has not been reported previously, and it is not present in the gnomAD browser; Polyphen-2, SIFT, and CADD score classified this variant as pathogenic.…”

“…1b). Among them, the p.I188N, p.G221R, p.A224V, p.G276R, and p.G401R mutations have previously been reported to be pathogenic [15,[19][20][21][22]. Four patients inherited their mutations from their mothers.…”