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. Author manuscript; available in PMC: 2017 Aug 31.
Published in final edited form as: Nat Commun. 2014;5:3301. doi: 10.1038/ncomms4301

Figure 4. Characterization of targets that affect copper levels in HeLa cells.

Figure 4

(a) Effect of targets down-regulation on ATP7A localization in basal conditions. Following gene-specific siRNA mediated knockdowns, HeLa cells were fixed, stained with the antibodies against ATP7A (green) and the trans-Golgi network marker TGN46 (red) and analyzed by confocal microscopy. The merged image is shown in the right panel. Scale bar: 25 µm. (b) Effect of target down-regulation on ATP7A protein levels and electrophoretic mobility. Following gene knockdowns, HeLa cell lysates were analyzed by Western blotting. The blot was cut at 130 kDa; the upper part was immunostained for ATP7A and the lower part was immunostained for Na/K-ATPase. A representative blot is shown; the entire blot is included as Supplementary Figure 6. (c) Quantitation of Western blot data by densitometry. The ATP7A protein levels were normalized to Na/K-ATPase protein levels and the values were compared to the intensity of ATP7A in cells transfected with a non-targeting siRNA (scrRNA). The error bars correspond to standard deviations values from four independent experiments. Asterisks mark significant changes (Mann-Whitney U Test, p<0.05) (lower panel).