Generation of NIS mutants with altered substrate specificity and Na+/Anion stoichiometry: potential therapeutic implications for NIS gene transfer

Abstract

Active I- transport into the thyroid gland for thyroid hormone biosynthesis is mediated by the Na+/I - symporter (NIS). I- uptake by NIS is electrogenic: 2 Na+ are translocated with each I- . I- transport defect (ITD) is an autosomal recessive condition caused by mutations in NIS which results in congenital hypothyroidism. We have obtained mechanistic information on NIS by characterizing G93R NIS, identified in a patient with congenital hypothyroidism due to ITD. G93R NIS is targeted to the plasma membrane but is inactive. Amino acid substitutions at position 93 show that the longer the side chain of the neutral residue at this position, the higher the Km for the anion substrates. Unlike WT NIS, which mediates symport of Na+ and the environmental pollutant perchlorate electroneutrally, G93T/N/Q/E/D NIS, strikingly, do so electrogenically with a 2:1 stoichiometry. Furthermore, G93E/Q NIS discriminate between anion substrates. We have generated a 3D homology model for NIS based on the Na+/galactose transporter (vSGLT) from Vibrio parahaemolyticus, a homologue of the eukaryotic Na+/glucose transporter (SGLT-1), and proposed a mechanism in which changes from an outwardly to an inwardly open conformation during the transport cycle use G93 as a pivot. Thus, the nature of the side chain at position 93 of NIS may control the extent as well as the final state of the transition between the outwardly and the inwardly open conformations and play a role in the kinetic parameters and the stoichiometry of the protein.;The ability of the thyroid to accumulate I- via NIS has been used for >60 years to eliminate surgical remnants of thyroid cancer and its metastases with 131I. Gene transfer can be used to induce NIS expression in tumors using tissue-specific promoters. We sought to use NIS-mediated 131I uptake to inhibit angiogenesis and tumor growth by targeting Tie2-expressing monocytes, a subset of cells that home to sites of angiogenesis. While TEMs are a promising way of using NIS to target angiogenesis, further studies are required to investigate the attractive possibility of using NIS in this manner to inhibit tumor growth.