Submitted on May 26, 2006
Accepted on September 15, 2006
Cell-based imaging of sodium iodide symporter activity with the yellow fluorescent protein variant YFP-H148Q/I152L
Kerry J Rhoden1*, Stefano Cianchetta1, Valeria Stivani1, Carla Portulano1, Luis Galietta2, and Giovanni Romeo1
1 UO Genetica Medica, University of Bologna, Bologna, Italy
2 Laboraty of Molecular Genetics, Istituto Giannina Gaslini, Genova, Italy
* To whom correspondence should be addressed. E-mail: kerry.rhoden{at}med.unibo.it.
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 (i) high affinity for extracellular I- (35 µM), (ii) inhibition by the NIS inhibitor perchlorate, (iii) extracellular Na+-dependence and (iv) 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 single cell differences 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 non-thyroidal cells following gene transfer, and suggest that fluorescence detection of cellular I- may be a useful tool to study the pathophysiology and pharmacology of NIS.
