MuRF1-mediated TRα nuclear accumulation following T3 treatment is dependent on TRα ligand-binding domain (E/F region) lysines. HL-1 cardiomyocytes were transfected with plasmids containing FLAG-TRα with various lysine to arginine (KR) mutants (Supplementary Fig. S2), transduced with AdMuRF1 for 4 h, and treated with 16.6 nM T3 for 2 h. (A) Nuclear localization of TRα was assessed using confocal immunofluorescent imaging of TRα (anti-FLAG, red) and nuclei (DAPI, blue). (B) MuRF1 (anti-myc, green) and TRα at the nucleus was assessed TRα (anti-FLAG, red). (C) TRα (anti-FLAG, red) and endogenous CAP350 (green). Data represent the mean ± S.E.M. A one-way ANOVA test was used to determine statistical significance. ##P < 0.001 as determined by a pairwise post-test, 400× final magnification.