Structure/function analysis of the sodium/iodine symporter (NIS) through characterization of congenital iodide transport defect (ITD)-causing NIS mutations

Abstract

Iodide (I-) is an essential component of the thyroid hormones T3 and T4, which are required for development of the central nervous system and intermediary metabolism in all tissues. I - is transported into thyroid epithelial cells via the Na+ /I- symporter (NIS). The cDNA encoding NIS was first identified in our laboratory, where an extensive structure/function characterization of NIS is being carried out. The diagnostic and therapeutic use of radioiodide in gene transfer experiments is fully dependent on NIS activity.;Several NIS mutants have been identified as a cause of congenital I - transport defect (ITD), including V59E NIS. ITD is characterized by low thyroid I- uptake, hypothyroidism, goiter, and a low saliva/plasma I- ratio. An early report on V59E NIS by Fujiwara et al.1 contended that V59E NIS exhibited ∼30% of the activity of wild-type NIS. However, this observation was at variance with the patients' phenotype of total lack of activity. We thoroughly characterized the V59E NIS mutation and studied the effects of several substitutions at position 59 of NIS. Each resulted in proper expression and plasma membrane targeting of NIS. We demonstrated that, contrary to Fujiwara et al.'s contention, V59E NIS is inactive. Indeed, E and all other charged amino acids or P at position 59 yielded inactive NIS proteins. In contrast, I- uptake was rescued to different degrees by the other substitutions. In summary, the structural requirement for NIS function at position 59 is a neutral, helix-promoting amino acid.;Another ITD-causing NIS mutation is T354P, which was initially reported to be inactive due to a disruption of transmembrane segment (TMS) IX by P. We have demonstrated that proper NIS function depends on the presence of an amino acid with a beta-hydroxyl group (S/T) at position 354. We have determined that T351, S353, T354, S356 and T357 of NIS TMS IX are important for NIS function and may play a role in the Na+ translocation pathway.;We also showed that, surprisingly, the 3 charged residues (D16, E79, and R208) that reside within predicted NIS TMS plays a role in I- transport. However, we identified, for the first time, NIS molecules (R208A and R208C NIS) with a decreased Km (higher apparent affinity) for Na+, a finding with potential clinical relevance, particularly in gene transfer experiments.;1Fujiwara H, Tatsumi K, Tanaka S, Kimura M, Nose O, Amino N 2000 A novel V59E missense mutation in the sodium iodide symporter gene in a family with iodide transport defect. Thyroid 10:471-4.