Fig. 4. (A) Schematic of the different mutant versions of gro transgenes. The hatched box in Gro
WD40 corresponds to 21 foreign amino acids added as a consequence of the construction strategy. (B-G) S2 cells transfected with different gro mutant expression vectors and stained with the anti-Gro (red – all panels except D) or pan-TLE monoclonal antibody (red – D) and TOPROIII (blue – overlap with Gro immunostaining seen as magenta) as a DNA counterstain – co-transfection with GFP was used to identify the transfected cells (not shown). (B) Grocdc2-, (C) GroQ, (D) GroGCS, (E) GroNLS- and (F,G) GroWD40. Note that transfected Gro is overexpressed with respect to endogenous; G is the same panel as F, except that the detection sensitivity is increased for red and endogenous Gro immunoreactivity is detectable in the three untransfected cells (arrows). As exogenous Gro is in excess, the localization observed is not influenced by interaction with endogenous Gro. Wild-type Gro also accumulates in the nucleus: in H, UAS-gro+ (red) was co-transfected with HA-tagged full length ci and counterstained with GFP (green), which localizes predominantly in the nucleus, but also in the cytoplasm. I shows another cell from the same transfection stained with an anti-HA antibody to detect Ci (red), which is exclusively cytoplasmic, thus providing no evidence for Ci-Gro interaction.
