FIGURE 9:

Loss of SDPN-1 disrupts association of filamentous actin with early endosomes. All micrographs are from deconvolved 3D confocal image stacks acquired in intact living animals expressing intestinal-specific LifeAct-GFP and tagRFP-RAB-5 in control RNAi (A–A′′′) and sdpn-1(RNAi) (B–B′′′). In control animals, many tagRFP::RAB-5-positive endosomes are positive for LifeAct::GFP. White arrowheads indicate positive colocalization. (B–B′′′) A striking decrease in localization of LifeAct::GFP and tagRFP::RAB-5 was seen in sdpn-1 RNAi animals. Gray arrows indicate early endosomes depleted of filamentous actin. In each image, autofluorescent lysosome-like organelles appear in in all three channels (including blue), whereas GFP appears only in the green channel and RFP appears only in the red channel. Green and red signals that do not overlap with the blue channel represent pure GFP and RFP signals, respectively. (C) Pearson’s r for colocalization of LifeAct::GFP with tagRFP::RAB-5. Six animals. Error bars represent SEM.***p < 0.001 (Student’s t test) Scale bar, 10 μm.