Mutations of the KCNQ1 and KCNE1 genes have been identified in individuals suffering from the hereditary cardiac long QT (LQT) syndromes. KCNQ1 (pore-forming subunit) and KCNE1 (accessory subunit) encode the potassium channel proteins and mutations in these subunits can lead to a loss or decrease in repolarising current, I_Ks, via an unknown mechanism. The aim of this study was to examine the effect known KCNE1 mutations (which cause LQT-5) had on the trafficking of the pore-forming subunit, KCNQ1. The C-terminus of the long isoform of KCNQ1 was tagged with enhanced green fluorescent protein (EGFP) and transiently transfected either alone; with wild type KCNE1 or mutant KCNE1 into CHO-K1 cells. ER co-localisation was determined by co-transfection of pDsRed2-ER (an ER-retained red fluorescent protein, Clontech, UK). Images were acquired with a Bio-Rad Radiance 2000 scanning laser confocal microscope. Co-localisation of green and red fluorescence was quantified using LaserPix software (Bio-Rad, UK). Data are expressed as means±SEM. Statistical analysis was performed using Student’s unpaired t-test or one-way ANOVA, as appropriate. Expression of wild type (WT) KCNQ1-GFP alone resulted in 38.3±2% (n=66) of the protein being ER retained. Co-expression with WT KCNE1 resulted in a reduction of ER retention to 21.6±2% (n=53, P<0.001). Co-expression of KCNQ1 with five of the KCNE1 mutations (G52R, T58P/L59P, S74L, D76N and R98W) did not significantly affect the export of KCNQ1 from the ER in comparison to the WT KCNQ1 control alone (range 34-39%, n=36-46). In contrast, a sixth mutation, T71I, did significantly (P<0.001) reduce the degree of ER retention to a level similar to that of the KCNQ1+WT KCNE1 control (21.0±2% n=49). As five of the mutations did not appear to interact with KCNQ1 in the ER we then investigated whether these mutations had a dominant-negative effect on trafficking of WT KCNQ1 when co-expressed with WT KCNE1. Four of the mutations (T71I , G52R, T58P/L59P and S74L) did not produce significantly different degrees of ER retention compared to the WT controls (range 19-22%, n=14-36). However, two of the mutations (D76N and R98W) did appear to have a significant dominant-negative affect on ER retention (range 36-44%, n=13-23, P<0.001) when compared to the WT controls. The results from this study suggest that LQT5 mutations in KCNE1 can cause a variety of functional effects including a general failure to promote ER export of the channel complex and a dominant-negative effect on the trafficking of the wild-type channel.