RAF1 kinase contributes to autophagic lysosome reformation

Autophagic lysosome reformation (ALR) is crucial for lysosomal homeostasis and therefore for different autophagic processes. Despite recent advances, the signaling mechanisms regulating ALR are incompletely understood. We show that RAF1, a member of the RAS/RAF/MEK/ERK pathway initiated by growth factors, has an essential, kinase-dependent role in lysosomal biology. RAF1 ablation impairs autophagy, and a proxisome screen identifies several proteins involved in autophagic and lysosomal pathways in the RAF1 molecular space. Two of these, SPG11 and the lipid phosphatase MTMR4, are RAF1 substrates. RAF1 ablation causes the appearance of enlarged autolysosomes and alters the phosphoinositide composition of autolysosomes. RAF1 and MTMR4 colocalize on autolysosomes, and overexpression of a MTMR4 mutant mimicking phosphorylation of the RAF1-dependent site rescues the lysosomal phenotypes induced by RAF1 ablation. Our data identify an RAF1 function in lysosomal homeostasis and a substrate through which the kinase regulates phospholipid metabolism at the lysosome, ALR, and autophagy.