Lysosomes respond to environmental cues by controlling their own biogenesis, but the underlying mechanisms are poorly understood. Here we describe a protein kinase C (PKC)-dependent and mTORC1-independent mechanism for regulating lysosome biogenesis, which provides insights into previously reported effects of PKC on lysosomes. By identifying lysosome-inducing compounds we show that PKC couples activation of the TFEB transcription factor with inactivation of the ZKSCAN3 transcriptional repressor through two parallel signalling cascades. Activated PKC inactivates GSK3β, leading to reduced phosphorylation, nuclear translocation and activation of TFEB, while PKC activates JNK and p38 MAPK, which phosphorylate ZKSCAN3, leading to its inactivation by translocation out of the nucleus. PKC activation may therefore mediate lysosomal adaptation to many extracellular cues. PKC activators facilitate clearance of aggregated proteins and lipid droplets in cell models and ameliorate amyloid β plaque formation in APP/PS1 mouse brains. Thus, PKC activators are viable treatment options for lysosome-related disorders.

The cell surface receptor CED-1 mediates apoptotic cell recognition by phagocytic cells, enabling cell corpse clearance in Caenorhabditis elegans. Here, we found that the C. elegans intracellular protein sorting complex, retromer, was required for cell corpse clearance by mediating the recycling of CED-1. Retromer was recruited to the surfaces of phagosomes containing cell corpses, and its loss of function caused defective cell corpse removal. The retromer probably acted through direct interaction with CED-1 in the cell corpse recognition pathway. In the absence of retromer function, CED-1 associated with lysosomes and failed to recycle from phagosomes and cytosol to the plasma membrane. Thus, retromer is an essential mediator of apoptotic cell clearance by regulating phagocytic receptor(s) during cell corpse engulfment.

C. elegans SORF-1 and SORF-2 and their mammalian homologs WDR91 and WDR81 maintain appropriate PtdIns3P levels in early-to-late endosome conversion by forming a complex with the Beclin1 subunit of the PI3K complex.

Lysosomes are degradation and signaling organelles that adapt their biogenesis to meet many different cellular demands; however, it is unknown how lysosomes change their numbers for cell division. Here, we report that the cyclin-dependent kinases CDK4/6 regulate lysosome biogenesis during the cell cycle. Chemical or genetic inactivation of CDK4/6 increases lysosomal numbers by activating the lysosome and autophagy transcription factors TFEB and TFE3. CDK4/6 interact with and phosphorylate TFEB/TFE3 in the nucleus, thereby inactivating them by promoting their shuttling to the cytoplasm. During the cell cycle, lysosome numbers increase in S and G2/M phases when cyclin D turnover diminishes CDK4/6 activity. These findings not only uncover the molecular events that direct the nuclear export of TFEB/TFE3, but also suggest a mechanism that controls lysosome biogenesis in the cell cycle. CDK4/6 inhibitors promote autophagy and lysosome-dependent degradation, which has important implications for the therapy of cancer and lysosome-related disorders.

Early-to-late endosome conversion involves switching of early endosomes Rab5 and PtdIns3P to late endosomes Rab7 and PtdIns(3,5)P2. Liu et al. identify WDR91 as a Rab7 effector that couples Rab switching with PtdIns3P down-regulation on endosomes and show that WDR91 is essential for neuronal development.

Mutations in WDR81, a regulator of the endosomal–lysosomal pathway, are implicated in CAMRQ2 syndrome, which manifests as cerebellar ataxia, mental retardation, and quadrupedal locomotion in patients. In this study, Liu et al. uncover a distinct function of WDR81 in the clearance of ubiquitinated and aggregated proteins by autophagy.

Nucleophosmin (NPM) is a multifunctional protein involved in both proliferation and apoptosis. Importantly, NPM negatively regulates p53 and is frequently overexpressed in a wide variety of cancers. To identify inhibitory molecules of NPM, we used an in vitro selection method termed systematic evolution of ligands by exponential enrichment (SELEX) to select RNA aptamers that bind to NPM with high affinity and specificity. The selected RNA aptamers bind to the central acidic region of NPM and affect its oligomerization both in vitro and in vivo. Remarkably, expression of NPM-specific aptamers causes mislocalization of NPM in the nucleoplasm rather than in the nucleolus, suggesting that NPM oligomerization is important for its proper localization. Moreover, p14ARF is mislocalized in the nucleoplasm and p53 is upregulated in cells expressing NPM aptamers. In addition, cancer cells expressing NPM aptamers not only undergo apoptosis on their own, but are more susceptible to apoptosis induced by DNA-damaging agents as well. These results suggest that interfering with NPM oligomerization can inhibit NPM function and aptamers targeting NPM can serve as potential lead for developing anticancer drugs.

Cell type-specific modifications of conventional endosomal trafficking pathways lead to the formation of lysosome-related organelles (LROs). C. elegans gut granules are intestinally restricted LROs that coexist with conventional degradative lysosomes. The formation of gut granules requires the Rab32 family member GLO-1. We show that the loss of glo-1 leads to the mistrafficking of gut granule proteins but does not significantly alter conventional endolysosome biogenesis. GLO-3 directly binds to CCZ-1 and they both function to promote the gut granule association of GLO-1, strongly suggesting that together, GLO-3 and CCZ-1 activate GLO-1. We found that a point mutation in GLO-1 predicted to spontaneously activate, and function independently of it guanine nucleotide exchange factor (GEF), localizes to gut granules and partially restores gut granule protein localization in ccz-1(-) and glo-3(-) mutants. CCZ-1 forms a heterodimeric complex with SAND-1(MON1), which does not function in gut granule formation, to activate RAB-7 in trafficking pathways to conventional lysosomes. Therefore, our data suggest a model whereby the function of a Rab GEF can be altered by subunit exchange. glo-3(-) mutants, which retain low levels of GLO-3 activity, generate gut granules that lack GLO-1 and improperly accumulate RAB-7 in a SAND-1 dependent process. We show that GLO-1 and GLO-3 restrict the distribution of RAB-7 to conventional endolysosomes, providing insights into the segregation of pathways leading to conventional lysosomes and LROs.