CARBOXY-TERMINAL SIGNALS REGULATE SODIUM/IODIDE SYMPORTER TARGETING TO THE PLASMA MEMBRANE

NICOLA, JP; MARTIN, M

Rio de Janeiro

Congreso; XVI Congreso de la Sociedad Latinoamericana de Tiroides; 2017

Introduction: The sodium/iodide symporter (NIS), a 643 amino acid glycoprotein expressed at the plasma membrane ofthyroid follicular cells, mediates iodide accumulation for thyroid hormone biosynthesis and radioiodide transport for diagnosisand treatment of thyroid cancer. The cloning of NIS provided the basis to investigate the decrease in iodide accumulationin thyroid cancer relative to healthy thyroid cells. Instead of finding only the expected lower NIS expression, the majority ofthyroid cancers showed a surprisingly NIS overexpression as compared to the surrounding normal tissue but retained intracellularly.Therefore, it is of considerable clinical relevance to elucidate the mechanisms underlying NIS plasma membrane targeting,a pursuit that could lead to new therapeutic interventions to increase the effectiveness of radioiodide therapy. Objective:Short linear motifs have been shown to participate in protein sorting along the secretory pathway. Therefore, we investigatedthe role of these motifs in NIS transport to the plasma membrane under physiological conditions. Material and methods:Short linear motifs on human NIS carboxy terminus were identified using in silico computational analysis. We then performedsite-directed mutagenesis to disrupt these motifs and functional in vitro studies were performed on MDCK-II cells. Resultsand conclusions: The NIS mutant lacking the carboxy-terminus (Δ546-643) is intracellularly retained in MDCK-II cells, thussuggesting that its carboxy-terminus may act as a determinant in anchoring adaptor required for NIS sorting to the plasmamembrane. In silico analysis revealed different putative short linear motifs frequently involved in plasma membrane proteinstargeting to the cell surface. Therefore, we generated deletion mutants of the carboxy-terminus (Δ546-598, Δ578-618, Δ618-634, Δ634-639, Δ640-643) and site-directed mutants of putative sorting motifs (V580A, L583A, L587A, V588A, L594A,L612A, E621A). Functional evaluation revealed that the NIS mutant Δ546-598 containing a putative tryptophan-acidic motif(W565D566) is intracellularly retained. Moreover, we demonstrated that the PDZ binding-motif T640NL643 is not necessaryfor NIS sorting to the plasma membrane. Although the molecular mechanisms that determine NIS plasma membrane targetingin thyroid cells remain elusive, here, we provide evidence that the carboxy-terminus contains crucial information for NISfunctional cell surface expression.