Modeling the Biochemical Phenotype of MCT8 Mutations In Vitro: Resolving a Troubling Inconsistency

Allan-Herndon-Dudley syndrome (AHDS) is a severe genetic disease caused by mutations in the monocarboxylate transporter 8 (MCT8) gene. MCT8 mediates transport of thyroid hormones in and out of cells, which is thought to play a pivotal role for embryonic and postnatal development of the human brain. Disconcertingly, MCT8R271H leads to a severe form of AHDS but shows residual transport activity when expressed in several types of cultured cells. Here we try to determine the mechanism behind the transport function of MCT8R271H found in overexpressing cell systems. Mutations of Arg271 were introduced into human MCT8 and stably transfected into Madin-Darby canine kidney cells and the human-derived cell line JEG1. Radioactive thyroid hormone-uptake experiments were performed to analyze the pH-dependent effect of the mutation on transport activity. Arg271His transports thyroid hormones in and out of cells in a pH-dependent manner. Its transport activity increases below pH 7.3 and is clearly diminished at physiological pH. The Michaelis constant of the mutant is unaltered, whereas the maximum velocity is reduced. The expression of Arg271His in JEG1 cells leads to an almost nonfunctional transporter at physiological pH replicating the human phenotype for this mutant in vitro and demonstrates, again, that mutant MCT8 activity depends on cellular background. The protonation of His271 at acidic pH restores activity of the mutant protein, which is not active in its deprotonated form at physiological pH. Thus, experimental parameters must be controlled carefully when modeling MCT8 deficiency in cells.