Autophagosome–lysosome fusion is independent of V-ATPase-mediated acidification

Abstract

The ATP-dependent proton pump V-ATPase ensures low intralysosomal pH, which is essential for lysosomal hydrolase activity. Based on studies with the V-ATPase inhibitor BafilomycinA1, lysosomal acidification is also thought to be required for fusion with incoming vesicles from the autophagic and endocytic pathways. Here we show that loss of V-ATPase subunits in the Drosophila fat body causes an accumulation of non-functional lysosomes, leading to a block in autophagic flux. However, V-ATPase-deficient lysosomes remain competent to fuse with autophagosomes and endosomes, resulting in a time-dependent formation of giant autolysosomes. In contrast, BafilomycinA1 prevents autophagosome–lysosome fusion in these cells, and this defect is phenocopied by depletion of the Ca ²⁺ pump SERCA, a secondary target of this drug. Moreover, activation of SERCA promotes fusion in a BafilomycinA1-sensitive manner. Collectively, our results indicate that lysosomal acidification is not a prerequisite for fusion, and that BafilomycinA1 inhibits fusion independent of its effect on lysosomal pH.

Abstract

BafilomycinA1 is an autophagy inhibitor, presumably owing to its blocking effect on the lysosomal proton pump V-ATPase. Here the authors show that V-ATPase-deficient lysosomes can still fuse with autophagosomes, showing that lysosomal acidification and fusion are two separable, independent events.