The Q267E mutation in the sodium/iodide symporter (NIS) causes congenital iodide transport defect (ITD) by decreasing the NIS turnover number

Abstract

The Na+/I- symporter (NIS) is a key plasma membrane glycoprotein that mediates active iodide (I-) transport in the thyroid and other tissues. Since isolation of the cDNA encoding NIS (G. Dai, O. Levy, and N. Carrasco (1996) Nature 379, 458-460), ten mutations in NIS have been identified as causes of congenital iodide transport defect (ITD). Two of these mutations (T354P and G395R) have been thoroughly characterized at the molecular level. Both mutant NIS proteins are inactive but normally expressed and correctly targeted to the plasma membrane. The hydroxyl group at the β-carbon of residue 354 is essential for NIS function, whereas the presence of a charged or large side-chain at position 395 interferes with NIS function. We report the extensive molecular analysis of the Q267E mutation in COS-7 cells transfected with rat or human Q267E NIS cDNA constructs. We used site-directed mutagenesis to engineer various residue substitutions into position 267. In contrast to previous suggestions that Q267E NIS was inactive, possibly because of a trafficking defect, we conclusively show that Q267E NIS is modestly active and properly targeted to the plasma membrane. Q267E NIS exhibited lower Vmax values for I- than wild-type NIS, suggesting that the decreased level of activity of Q267E NIS is due to a lower catalytic rate. That Q267E NIS retains even partial activity sets this ITD-causing mutant apart from T354P and G395R NIS. The presence of charged residues (of any polarity) other than Glu at position 267 rendered NIS inactive without affecting its expression or targeting, but substitution with neutral residues at this position was compatible with partial activity.

Mechanism of anion selectivity and stoichiometry of the Na + /I - symporter (NIS)

“…1 A, D, and G and 4A). In contrast to the many NIS mutants we have studied previously (13)(14)(15)(16)(17)19), substitutions at position 93 show a significant change in the K m for I − . This is true for not only the neutral residues Thr, Asn, and Gln (Fig.…”

“…S1). The mutants that have been studied in detail have provided key mechanistic information on NIS (13)(14)(15)(16)(17). For example, substitutions at T354 have revealed that this position requires an -OH group at the β-carbon, which we showed is involved in Na þ binding/translocation, and proposed a structural homology between the Aquifex aeolicus Na þ ∕leucine transporter (LeuT) (18) and NIS, despite a lack of sequence homology (19).…”

Na + coordination at the Na2 site of the Na + /I − symporter

“…Biotinylation of cell surface proteins was performed as previously described (24). Briefly, transfected cells were incubated with 1 mg/mL of the membrane-impermeable biotin reagent sulfo-NHS-SS-biotin (Pierce Chemical), which covalently interacts with extracellular primary amines.…”

“…autosomal recessive condition that results from NIS being either inactive (22)(23)(24) or not targeted to the plasma membrane (16,17,25,26). The resulting impairment in I − uptake in the thyroid leads to congenital hypothyroidism, which, unless treated early in life by TH administration, causes goiter and even mental retardation.…”

The importance of sodium/iodide symporter (NIS) for thyroid cancer management

“…Congenital hypothyroidism due to impaired iodide uptake has long been described in the literature; however, NIS mutations were recognized only after the cloning of NIS cDNA, and correspond to point mutations [G93R, Q267E, C272X, T354P, Y531X, G543E, G395R, V59E or frameshift (515X)]. Functional studies have already been done for some mutated NIS and have confirmed the following changes in NIS molecular characteristics: loss of function of the protein (T354P, G395R), partial activity (Q267E) or impaired targeting of the protein to the plasma membrane (G543E) (81,82).…”