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Am J Physiol Cell Physiol 292: C814-C823, 2007. First published September 20, 2006; doi:10.1152/ajpcell.00291.2006 Free Article
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METHODS IN CELL PHYSIOLOGY

Cell-based imaging of sodium iodide symporter activity with the yellow fluorescent protein variant YFP-H148Q/I152L

Kerry J. Rhoden,1 Stefano Cianchetta,1 Valeria Stivani,1 Carla Portulano,1 Luis J. V. Galietta,2 and Giovanni Romeo1

1Laboratory of Medical Genetics, Department of Internal Medicine, Cardiology and Hepatology, University of Bologna, Bologna; and 2Laboratory of Molecular Genetics, Istituto Giannina Gaslini, Genova, Italy

Submitted 26 May 2006 ; accepted in final form 15 September 2006

The sodium iodide symporter (NIS) mediates iodide (I–) transport in the thyroid gland and other tissues and is of increasing importance as a therapeutic target and nuclear imaging reporter. NIS activity in vitro is currently measured with radiotracers and electrophysiological techniques. We report on the development of a novel live cell imaging assay of NIS activity using the I–-sensitive and genetically encodable yellow fluorescent protein (YFP) variant YFP-H148Q/I152L. In FRTL-5 thyrocytes stably expressing YFP-H148Q/I152L, I– induced a rapid and reversible decrease in cellular fluorescence characterized by 1) high affinity for extracellular I– (35 µM), 2) inhibition by the NIS inhibitor perchlorate, 3) extracellular Na+ dependence, and 4) TSH dependence, suggesting that fluorescence changes are due to I– influx via NIS. Individual cells within a population of FRTL-5 cells exhibited a 3.5-fold variation in the rate of NIS-mediated I– influx, illustrating the utility of YFP-H148Q/I152L to detect cell-to-cell difference in NIS activity. I– also caused a perchlorate-sensitive decrease in YFP-H148Q/I152L fluorescence in COS-7 cells expressing NIS but not in cells lacking NIS. These results demonstrate that YFP-H148Q/I152L is a sensitive biosensor of NIS-mediated I– uptake in thyroid cells and in nonthyroidal cells following gene transfer and suggest that fluorescence detection of cellular I– may be a useful tool by which to study the pathophysiology and pharmacology of NIS.

thyroid; fluorescence microscopy; FRTL-5 cells


Address for reprint requests and other correspondence: K. J. Rhoden, U. O. Genetica Medica, Padiglione 11, Policlinico S. Orsola-Malpighi, via Massarenti 9, 40138 Bologna, Italy (e-mail: kerry.rhoden{at}med.unibo.it)




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