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. Author manuscript; available in PMC: 2009 Nov 28.
Published in final edited form as: Cell. 2008 Nov 28;135(5):838–851. doi: 10.1016/j.cell.2008.09.041

FIGURE 3. Disruption of autophagy in neural tissues in trpml.

FIGURE 3

(A–D) Transmission EMs of 21 day-old photoreceptor cells: (A and B) wt, (C and D) trpml1. (B and D) Magnifications of the boxed regions in (A and C). The red and yellow arrows in (D) indicate multilamellar and multivesicular bodies respectively.

(E) Fold-increase in autophagosome-like vesicles in trpml1 photoreceptors. n=5 animals, ≥30 ommatidia per animal; *, p≤10−4.

(F and G) Brains (21 day-old flies) expressing GFP-ATG8 and viewed at 488 nm: (F) wt, (G) trpml1.

(H) Fold-increase in GFP-ATG8 fluorescence in trpml1 brains. n=3; *, p≤5×10−4.

(I–N) Ommatidia from 21 day-old flies expressing GFP-ATG8: (I–K) control, (L–N) trpml1. Ommatidia were viewed at 488 nm to detect GFP-ATG8 (I–L) and 568 nm to detect LysoTracker (Lyso; J–M). (K and N) Merges.

(O) Fold-increase in GFP-positive vesicles in trpml1 ommatidia. n=3 independent experiments; *, p≤5×10−5.

(P) Rh1 remaining after 6 h exposure to blue light (480 nm) in 5 day-old flies. Means are relative to the starting value (defined as 100%). n=3; *, p≤0.05.

(Q and R) 4 week-old brains stained with anti-ubiquitin antibodies (Supplemental Experimental Procedures): (Q) wt, (R) trpml1.

(S) Western blot of head extracts (4 week-old flies) probed with anti-ubiquitin antibodies and reprobed with anti-Tubulin antibodies.

(T) Fold increase of ubiquitination in head extracts based on Western blotting. Data were normalized to wt (1.0). n=3; *, p ≤0.05.

(U) Mitochondria per photoreceptor cell in 21 day-old wt flies. n=5 animals; *, p≤0.05.

(V and W) Ommatidia from 21 day-old wt (V) and trpml1 (W) flies viewed at 310 nm to detect DAPI (blue) and 568 nm to detect MitoTracker-orange CM-H2TMRos (red).

(X) Fold decrease in MitoTracker-orange CM-H2TMRos staining in trpml1 head extracts, normalized to wt (set at 1.0). n=3; *, p≤5×10−6.

(Y) Relative H2O2 levels in whole fly extracts. Data were normalized to wt (set at 1.0). n≥6, 10 flies/experiment; *, p≤5×10−6. Dissected brains and ommatidia were viewed by confocal microscopy.

All statistical analyses, t-test.