title: Vps33B is required for delivery of endocytosed cargo to lysosomes.
abstract: Lysosomes are the main degradative compartments of eukaryotic cells. The CORVET and HOPS tethering complexes are well known for their role in membrane fusion in the yeast endocytic pathway. Yeast Vps33p is part of both complexes, and has two mammalian homologues: Vps33A and Vps33B. Vps33B is required for recycling of apical proteins in polarized cells and a causative gene for ARC syndrome. Here, we investigate whether Vps33B is also required in the degradative pathway. By fluorescence and electron microscopy we show that Vps33B depletion in HeLa cells leads to significantly increased numbers of late endosomes that together with lysosomes accumulate in the perinuclear region. Degradation of endocytosed cargo is impaired in these cells. By electron microscopy we show that endocytosed BSA-gold reaches late endosomes, but is decreased in lysosomes. The increase in late endosome numbers and the lack of internalized cargo in lysosomes are indicative for a defect in late endosomal-lysosomal fusion events, which explains the observed decrease in cargo degradation. A corresponding phenotype was found after Vps33A knock down, which in addition also resulted in decreased lysosome numbers. We conclude that Vps33B, in addition to its role in endosomal recycling, is required for late endosomal-lysosomal fusion events.
authors.affiliation: Department of Cell Biology and Institute of Biomembranes, Center for Molecular Medicine, University Medical Center Utrecht, Heidelberglaan 100, 3584CX, Utrecht, The Netherlands.|Present address: Institut Jacques Monod, CNRS, UMR7592, Université Paris Diderot, Sorbonne Paris Cité, F-75013, Paris, France.
authors.forename: Romain
authors.initials: R
authors.lastname: Galmes
authors.affiliation: Department of Cell Biology and Institute of Biomembranes, Center for Molecular Medicine, University Medical Center Utrecht, Heidelberglaan 100, 3584CX, Utrecht, The Netherlands.
authors.forename: Corlinda
authors.initials: C
authors.lastname: ten Brink
authors.affiliation: Department of Cell Biology and Institute of Biomembranes, Center for Molecular Medicine, University Medical Center Utrecht, Heidelberglaan 100, 3584CX, Utrecht, The Netherlands.|Present address: Monash Micro Imaging, 15 Innovation Walk, Strip 1 Monash Biotechnology, Monash University, Clayton, VIC, 3800, Australia.
authors.forename: Viola
authors.initials: V
authors.lastname: Oorschot
authors.affiliation: Department of Cell Biology and Institute of Biomembranes, Center for Molecular Medicine, University Medical Center Utrecht, Heidelberglaan 100, 3584CX, Utrecht, The Netherlands.
authors.forename: Tineke
authors.initials: T
authors.lastname: Veenendaal
authors.affiliation: Department of Cell Biology and Institute of Biomembranes, Center for Molecular Medicine, University Medical Center Utrecht, Heidelberglaan 100, 3584CX, Utrecht, The Netherlands.
authors.forename: Caspar
authors.initials: C
authors.lastname: Jonker
authors.affiliation: Department of Cell Biology and Institute of Biomembranes, Center for Molecular Medicine, University Medical Center Utrecht, Heidelberglaan 100, 3584CX, Utrecht, The Netherlands.
authors.forename: Peter
authors.initials: P
authors.lastname: van der Sluijs
authors.affiliation: Department of Cell Biology and Institute of Biomembranes, Center for Molecular Medicine, University Medical Center Utrecht, Heidelberglaan 100, 3584CX, Utrecht, The Netherlands.
authors.forename: Judith
authors.initials: J
authors.lastname: Klumperman
mesh_terms: D004705:Endocytosis / Q000502:physiology*; D011992:Endosomes / Q000378:metabolism* / Q000648:ultrastructure; D055785:Gene Knockdown Techniques; D006367:HeLa Cells; D006801:Humans; D008247:Lysosomes / Q000378:metabolism* / Q000648:ultrastructure; D008561:Membrane Fusion / Q000502:physiology; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D021381:Protein Transport; D033921:Vesicular Transport Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: C411584:VPS33B protein, human; D033921:Vesicular Transport Proteins
keywords: HOPS complex; SM proteins; Vps33A; Vps33B; late endosomes; lysosomes