Characterization of two types of CyPrPEGFP aggregates in different cells. (A) The cellular localization of CyPrPEGFP and GFP-250 was visualized after 24 h of expression in N2a, Hela, Cos7, and Huh-7 cells. Nuclei were stained with Hoechst (blue). Green and blue channels are shown merged. (B) Western blot of CyPrPEGFP and GFP-250 in 100 μg of protein extract from detergent-soluble (Sup.) and -insoluble (Pellet) fractions of N2a and Hela cells. The percentage of protein in the supernatant and pellet indicated below each blot was measured by densitometric analysis, and represents the mean and S.D. of three independent experiments. (C) Western blot of CyPrPEGFP and GFP-250 in different fractions of a 1.2 to 2 M sucrose gradient loaded with lysates from N2a or Hela cells. This experiment is representative of two independent experiments. (D) The percentage of cells displaying aggresomes (black columns) or dispersed aggregates (white columns) was calculated in N2a, HEK293, Hela, and Huh-7 cells 24 h post-transfection. (E) The percentage of N2a (black columns) and Hela (white columns) cells displaying aggresomes or dispersed aggregates was calculated different times post-transfection with CyPrPEGFP or GFP-250. (D, E) Data represent the mean and S.D. of three independent experiments. More than 200 cells were counted for each condition.
Beaudoin et al. BMC Cell Biology 2008 9:59 doi:10.1186/1471-2121-9-59