The Retromer Complex Is Required for Rhodopsin Recycling and Its Loss Leads to Photoreceptor Degeneration
Figure 5
Rh1 levels are decreased in Vps26 mutant PRs after internalization.
(A) Western blots for Rh1 and TRP on control (iso) and Vps261 mutant PRs. Flies are kept in the dark, exposed to white light for 16 h to induce endocytosis and lysosomal degradation of Rh1, or exposed to 16-h white light and allowed to recover in the dark for 6 h. The dark-reared control and Vps261 mutants show similar Rh1 levels. Upon 16-h exposure to white light, Rh1 levels in Vps261 decrease more than those in control. After recovery in the dark, both control and Vps261 mutants exhibit comparable Rh1 levels. We loaded 0.4 fly heads in each lane. (B) Quantification of the levels of Rh1 normalized to TRP. Three independent experiments were performed. Student's t test; error bars represent SEM; * p<0.01; ns, no significance. (C) ERG traces of Vps261 mosaic flies kept in the dark, or exposed to light for 16 h and then recovered in the dark for 6 h. ERG responses of Vps261 mutant PRs are not impaired when exposed to these illumination paradigms. (D) Quantification of ERG amplitude in (C). ERGs of 10 flies were quantified for each condition. ERG amplitudes are normalized to dark-reared Vps261 mutant PRs. Error bars represent SEM; ns, no significance. (E) Rh1 colocalizes with Vps26 in control PRs. Rh1 internalization is induced by the pulse-chase protocol used in Figure 4B. Upon a 2-h recovery in the dark, control PRs are stained with Rh1 and Vps26 antibodies. Multiple Rh1 punctae colocalize with Vps26.