Bartter’s syndrome comprises a group of renal tubular disorders characterised by salt and fluid wasting and associated hypotension (Hebert et al. 2005). Mutations in the gene KCNJ1, which encodes the small conductance K⁺ channel ROMK (Kir 1.1), are responsible for Type II Bartter’s syndrome. ROMK mediates K⁺ recycling across the apical membranes of the thick ascending limb (TAL) and K⁺ secretion in the distal nephron. Characterisation of mutations which disrupt channel function and/or trafficking and targeting contribute much to the understanding of ROMK physiology. A previous study (Peters et al. 2003) indicated that the trafficking of four Bartter’s mutants (T71M, V72E, A306T and F325C) was disrupted in non-polarized HEK 293 cells. In the present study, we investigated the properties of these mutants in polarized MDCK II cells in order to determine whether they also disrupt targeting. The cDNAs encoding human ROMK1 and Bartter’s mutants had been previously N-terminally tagged with eGFP(Peters et al. 2003). Confluent, polarized MDCK II cells grown on permeable filter supports were transiently transfected using Lipofectamine 2000^TM with 1μg cDNA encoding wild-type ROMK1 or ROMK1 Bartter’s mutants. At 48h post transfection, the monolayers were fixed with methanol:acetone (7:3 v/v) and nuclei were counterstained with 5 μg/ml prodium iodide (red). Intracellular localisation of the eGFP-fusion proteins was determined by fluorescence confocal microscopy. Transfections were repeated between 3 and 5 times for each construct. As expected, WT-ROMK1 was predominantly targeted to the apical pole in polarized MDCK II cells. The T71M (N-terminus) and A306T (C-terminus) mutants targeted to the apical domain and the fluorescence distribution pattern was similar to WT. In contrast, the V72E and F325C mutants showed a fluorescence pattern consistent with intracellular localisation and there was no distinct expression at either membrane domain. These results indicate that single amino acid mutations in either the N- and C-termini can have profound effects on trafficking and targeting, and underscore the importance of both termini in the ability of ROMK to be targeted normally.