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The mechanistic target of rapamycin complex 1 (mTORC1) is a protein kinase complex that localizes to lysosomes to up-regulate anabolic processes and down-regulate autophagy. Although mTORC1 is known to be activated by lysosome positioning and by amino acid-stimulated production of phosphatidylinositol 3-phosphate (PtdIns3P) by the lipid kinase VPS34/PIK3C3, the mechanisms have been elusive. Here we present results that connect these seemingly unrelated pathways for mTORC1 activation. Amino acids stimulate recruitment of the PtdIns3P-binding protein FYCO1 to lysosomes and promote contacts between FYCO1 lysosomes and endoplasmic reticulum that contain the PtdIns3P effector Protrudin. Upon overexpression of Protrudin and FYCO1, mTORC1-positive lysosomes translocate to the cell periphery, thereby facilitating mTORC1 activation. This requires the ability of Protrudin to bind PtdIns3P. Conversely, upon VPS34 inhibition, or depletion of Protrudin or FYCO1, mTORC1-positive lysosomes cluster perinuclearly, accompanied by reduced mTORC1 activity under nutrient-rich conditions. Consequently, the transcription factor EB enters the nucleus, and autophagy is up-regulated. We conclude that PtdIns3P-dependent lysosome translocation to the cell periphery promotes mTORC1 activation., (© 2017 Hong et al.)

Lysosome integrity is essential for cell viability, and lesions in lysosome membranes are repaired by the ESCRT machinery. Here, we describe an additional mechanism for lysosome repair that is activated independently of ESCRT recruitment. Lipidomic analyses showed increases in lysosomal phosphatidylserine and cholesterol after damage. Electron microscopy demonstrated that lysosomal membrane damage is rapidly followed by the formation of contacts with the endoplasmic reticulum (ER), which depends on the ER proteins VAPA/B. The cholesterol‐binding protein ORP1L was recruited to damaged lysosomes, accompanied by cholesterol accumulation by a mechanism that required VAP–ORP1L interactions. The PtdIns 4‐kinase PI4K2A rapidly produced PtdIns4P on lysosomes upon damage, and knockout of PI4K2A inhibited damage‐induced accumulation of ORP1L and cholesterol and led to the failure of lysosomal membrane repair. The cholesterol–PtdIns4P transporter OSBP was also recruited upon damage, and its depletion caused lysosomal accumulation of PtdIns4P and resulted in cell death. We conclude that ER contacts are activated on damaged lysosomes in parallel to ESCRTs to provide lipids for membrane repair, and that PtdIns4P generation and removal are central in this response. Synopsis: Lysosome integrity is essential for cell survival, and ruptures in lysosome membranes are repaired by the ESCRT machinery. This study identifies an additional, ESCRT‐independent repair mechanism that involves cholesterol transfer mediated by contact sites between lysosomes and the endoplasmic reticulum (ER). Upon lysosome damage, the ER proteins VAPA/B form contacts between ER and lysosomes and recruit the cholesterol‐binding protein ORP1L.PI4K2A‐dependent accumulation of PtdIns4P on the damaged lysosomes is essential for ORP1L recruitment and lysosome repair.ORP1L induces cholesterol accumulation on damaged lysosomes.Depletion of the cholesterol–PtdIns4P transporter OSBP causes PtdIns4P hyperaccumulation on damaged lysosomes and impairs cell viability. [ABSTRACT FROM AUTHOR]

The article presents the discussion on endosome following the degradative pathway or fusing with the plasma membrane to release exosomes. Topics include body fluids such as blood and urine containing exosomes making them useful as biomarkers in cancer; and endosomes vary in the luminal pH, show distinct subcellular localizations, and contain specific proteins and lipids.

Lysosomes fuse with the plasma membrane to help repair membrane lesions, but how they are positioned close to these lesions is not fully understood. Now, Encarnação et al. (2016. J. Cell Biol. http://dx.doi.org/10.1083/jcb.201511093) demonstrate that the lysosomal GTPase Rab3a and its effectors orchestrate lysosome positioning and plasma membrane repair. [ABSTRACT FROM AUTHOR]

Abstract: Clinical features characterizing Angelman syndrome, previously shown to be caused by disruption of UBE3A, were recently also described in neurologically disabled patients with mutations in SLC9A6, which encodes the Na+/H+ exchanger NHE6. In the present work we have focused on NHE6Δ255–256, the protein product of a specific 6-bp patient deletion in SLC9A6. To resolve the molecular mechanism causing the cellular dysfunction associated with this mutant, we have characterized its intracellular behaviour in comparison to wild type NHE6. Our study demonstrates that NHE6Δ255–256 is much less stable than the wild type protein. Whereas wild type NHE6 is transported to the plasma membrane and early endosomes and remains stable, NHE6Δ255–256 is degraded via two independent pathways mediated by proteasomes and lysosomes, respectively. Depletion of NHE6 had no detectable effect on endosomal pH, but co-depletion of NHE6 and the closely related NHE9 caused enhanced acidification of early endosomes. Our results suggest that NHE6 participates in regulation of endosomal pH and provides a cellular basis for understanding the loss of NHE6 function leading to a neurological phenotype resembling Angelman syndrome. [Copyright &y& Elsevier]

Selective trafficking of membrane proteins to lysosomes for destruction is required for proper cell signalling and metabolism. Ubiquitylation aids this process by specifying which proteins should be transported to the lysosome lumen by the multivesicular endosome pathway. The endosomal sorting complex required for transport (ESCRT) machinery sorts cargo labelled with ubiquitin into invaginations of endosome membranes. Then, through a highly conserved mechanism also used in cytokinesis and viral budding, it mediates the breaking off of the cargo-containing intraluminal vesicles from the perimeter membrane. The involvement of the ESCRT machinery in suppressing diseases such as cancer, neurodegeneration and infections underscores its importance to the cell. [ABSTRACT FROM AUTHOR]

Hrs has an essential role in sorting of monoubiquitinated receptors to multivesicular bodies for lysosomal degradation, through recognition of ubiquitinated receptors by its ubiquitin-interacting motif (UIM). Here, we present the structure of a complex of Hrs-UIM and ubiquitin at 1.7-Å resolution. Hrs-UIM forms a single α-helix, which binds two ubiquitin molecules, one on either side. These two ubiquitin molecules are related by pseudo two-fold screw symmetry along the helical axis of the UIM, corresponding to a shift by two residues on the UIM helix. Both ubiquitin molecules interact with the UIM in the same manner, using the Ile44 surface, with equal binding affinities. Mutational experiments show that both binding sites of Hrs-UIM are required for efficient degradative protein sorting. Hrs-UIM belongs to a new subclass of double-sided UIMs, in contrast to its yeast homolog Vps27p, which has two tandem single-sided UIMs. [ABSTRACT FROM AUTHOR]

Multivesicular endosomes are important as compartments for receptor downregulation and as intermediates in the formation of secretory lysosomes. Work during the past year has shed light on the molecular mechanisms of protein sorting into multivesicular endosomes and yielded information about the machinery involved in multivesicular endosome formation. Monoubiquitination functions as a signal for sorting transmembrane proteins into intraluminal vesicles of multivesicular endosomes and subsequent delivery to lysosomes. A molecular machinery that contains the ubiquitin-binding protein Hrs/Vps27 appears to be central in this sorting process. Three conserved multisubunit complexes, ESCRT-I, -II and -III, are essential for both sorting and multivesicular endosomes formation. Enveloped RNA viruses such as HIV can redirect these complexes from multivesicular endosomes to the plasma membrane to facilitate viral budding. [Copyright &y& Elsevier]

The mechanistic target of rapamycin complex 1 (mTORC1) is a protein kinase complex that localizes to lysosomes to up-regulate anabolic processes and down-regulate autophagy. Although mTORC1 is known to be activated by lysosome positioning and by amino acid-stimulated production of phosphatidylinositol 3-phosphate (PtdIns3P) by the lipid kinase VPS34/PIK3C3, the mechanisms have been elusive. Here we present results that connect these seemingly unrelated pathways for mTORC1 activation. Amino acids stimulate recruitment of the PtdIns3P-binding protein FYCO1 to lysosomes and promote contacts between FYCO1 lysosomes and endoplasmic reticulum that contain the PtdIns3P effector Protrudin. Upon overexpression of Protrudin and FYCO1, mTORC1-positive lysosomes translocate to the cell periphery, thereby facilitating mTORC1 activation. This requires the ability of Protrudin to bind PtdIns3P. Conversely, upon VPS34 inhibition, or depletion of Protrudin or FYCO1, mTORC1-positive lysosomes cluster perinuclearly, accompanied by reduced mTORC1 activity under nutrient-rich conditions. Consequently, the transcription factor EB enters the nucleus, and autophagy is up-regulated. We conclude that PtdIns3P-dependent lysosome translocation to the cell periphery promotes mTORC1 activation. [ABSTRACT FROM AUTHOR]