A New Paradigm for MAPK: Structural Interactions of hERK1 with Mitochondria in HeLa Cells (original) (raw)
Figure 4
hERK1 interaction with histones.
(A) HeLa cells transfected with hERK1-GFP, fixed and labelled with antibodies to H2A and H4 and a secondary antibody conjugated with Cy3. GFP and Cy3 images prior and after Cy3 photobleaching, a merged image of GFP and Cy3 prior to photobleaching and a FRET map in accord to Eq. 1 are shown. Bar = 10 µm. Color bar = FRET efficiency. Negative control: same FRET experiment in HeLa cells transfected with GFP and further stained against histones as above. Representative images are shown. (B) Colocalization indexes. M and C coefficients were estimated for both green and red images (Mgreen, Mred, Cgreen, Cred). P = Pearson's correlation coefficient, MO = Manders Overlap Coefficient. (C) Mean FRET efficiency estimated and analyzed as in Fig. 3; *p<0.5 with respect to GFP transfected cells. §p<0.5 with respect to hERK1-GFP transfected cells with no histone or Cy3 staining. (D) Pulldown experiment performed as in Fig. 2, with proteins detected by western blotting.