Basolateral Endocytic Recycling Requires RAB-10 and AMPH-1 Mediated Recruitment of RAB-5 GAP TBC-2 to Endosomes
Fig 7
Recycling cargo accumulates in RAB-5 labeled endosomes in tbc-2 and amph-1 mutants.
All images are from deconvolved 3D confocal image stacks acquired in intact living animals expressing a GFP-tagged recycling cargo protein, the IL-2 receptor alpha-chain (hTAC-GFP) and tagRFP-RAB-5 in wild-type animals (A-A‴), tbc-2(tm2241) (B-B‴) and amph-1(tm1060) mutants (C-C‴). (A-A‴) In wild-type animals, hTAC-GFP labels both punctate and tubular endosomal structures and has very little overlap with tagRFP-RAB-5. (B-B‴) In tbc-2 mutants, hTAC-GFP and tagRFP-RAB-5 label abnormally enlarged endosomal structures. There is a striking increase in the degree of overlap between hTAC-GFP and tagRFP-RAB-5 in tbc-2 mutants in comparison to that in wild-type animals. (C-C‴) In amph-1 mutants, the degree of colocalization between hTAC-GFP and tagRFP-RAB-5 also increased. In each image autofluorescent lysosome-like organelles appear in blue in all three channels, whereas GFP appears only in the green channel and RFP appears only in the red channel. Signals observed in the green or red channels that do not overlap with signals in the blue channel are considered bona fide GFP or RFP signals respectively. (Scale bar: 10 μm) (D) Pearson's correlation coefficient for colocalization of hTAC-GFP and tagRFP-RAB-5. n = 6. Error bars represent SEM. ***P<0.001(student's t test).
