Figure 4

Confocal microscopic analysis of Nedd4-2 and Fe65. Confocal microscopic analysis of transfected EGFP Nedd4-2 (wt) (A), EGFP Nedd4-2 AL mutant (B) (all constructs were shown as green color). The transfected EGFP-Nedd4-2 (wt) (detected in both the cytoplasm and the nucleus) was merged (yellow) with the endogenous Fe65 nuclear speckles around the nuclear pore (A). The transfected EGFP Nedd4-2 AL mutant was not detected in nuclear speckles around nuclear pore (B). The interaction between Fe65 and Nedd4-2 were detected as nuclear region around nuclear pore. However, Nedd4-2 AL mutant was not merged with Fe65 around nuclear pore (B, right lane). Confocal microscopic analysis of transfected EGFP-Fe65 (wt) and EGFP-Fe65 WW domain mutant (W243KP245K) (C). All constructs were shown as green color. The transfected EGFP-Fe65 (wt) (detected in both the cytoplasm and the nucleus) was merged (yellow) with the endogenous Nedd4-2 nuclear speckles around the nuclear pore (C). The transfected EGFP-Fe65 WW domain mutant was not detected in nuclear speckles around nuclear pore (D). The interaction between Fe65 and Nedd4-2 were detected as nuclear speckles around nuclear pore. EGFP-Fe65 WW domain mutant was not merged with Fe65 around nuclear pore (D, right lane).