Abstract

<p>Two lines of 3T3-L1 stably expressing wt FTO (hFTO-3 and hFTO-4) or R96Q (R96Q-2 and R96Q-4) were generated. (A) The protein expression level was confirmed by Western-blot using FTO and Myc antibodies, with β-actin as a loading control. (B) The sub-cellular localization of hFTO-4 or R96Q-4 was determined with confocal fluorescence imaging. Scale bars of 10μm are shown in the bottom row of images. (C) 3T3-L1differentiation was enhanced by over-expression of wt FTO and inhibited by R96Q. Level of differentiation was measured by extent of Oil Red-O staining at day 6 post differentiation induction (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0133788#sec002" target="_blank">materials and methods</a>). Demethylase activities of wt hFTO-4 and R96Q-4 in 3T3-L1 cells were determined by dot blotting (D) and quantified by Grayscale analysis with ImageJ software (E). Data are represented as means± SD of four replicates. * stands for p<0.05 and** stands for p<0.01in Student’s t-test.</p

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