A lysosomal delivery mechanism required for autophagosome degradation

Autophagy is a finely orchestrated cellular catabolic process through which cytoplasmic material is targeted for lysosomal degradation via a series of intracellular vesicle formation and fusion events. The final degradation step is essential for clearing autophagic cargo and recycling of macromolecules. We have identified a highly conserved, yet poorly characterised, small transmembrane protein named RNAseK as a novel regulator of autophagosome degradation. Analyses of RNAseK knockout cells revealed that, while autophagosome formation and maturation were intact, cargo degradation was severely disrupted. This phenotype was not a result of general lysosomal defect in the absence of RNAseK as lysosomal protease activity and acidification remained intact suggesting a specificity to autolysosome degradation. Comparison of lysosome preparations derived from wildtype or RNAseK knockout cells showed reduced levels of a subset of lysosomal hydrolases in the absence of RNAseK. Of these hydrolases, knockdown of PLD3 leads to a defect in autophagosome degradation. In addition, cells depleted of RNAseK exhibited an accumulation of the ESCRT-III complex component, VPS4a, in the lysosomal fraction, which is required for the lysosomal targeting of PLD3. Altogether, our findings identified a pathway required for hydrolase delivery to lysosomes in order to mediate efficient autolysosome degradation.