title: LYST affects lysosome size and quantity, but not trafficking or degradation through autophagy or endocytosis.
abstract: Mutations in the large BEACH domain-containing protein LYST causes Chediak-Higashi syndrome. The diagnostic hallmark is enlarged lysosomes and lysosome-related organelles in various cell types. Dysfunctional secretion of enlarged lysosome-related organelles has been observed in cells with mutations in LYST, but the capacity of the enlarged lysosomes to degrade endogenous proteins has not been studied. Here, we show for the first time that small interfering RNA-depletion of LYST in human cell lines recapitulates the LYST mutant phenotype of enlarged lysosomes. We found no evidence for an effect of LYST depletion on autophagy or endocytic degradation. Autophagosomes are formed in normal size and quantities and are able to fuse to the enlarged lysosomes, leading to normal rates of degradation. Degradation of the epidermal growth factor receptor (EGFR) was similarly not affected, indicating that the enlarged lysosomes are fully functional in degrading endogenous proteins. Retrograde trafficking of toxins as well as the localization of transporters of lysosomal proteins, adaptor protein-3 (AP-3) and cation-independent mannose-6-phosphate receptor (CI-MPR), were all found to be unaffected by LYST. Quantitative analysis of the enlarged lysosomes shows that LYST depletion causes a reduction in vesicle quantity per cell, while the total enzymatic content and vesicular pH are unaffected, supporting a role for LYST in lysosomal fission and/or fusion events.
authors.affiliation: Department of Biochemistry, Institute of Basic Medical Sciences, University of Oslo, PB 1112 Blindern, 0317 Oslo, Norway.
authors.forename: Petter
authors.initials: P
authors.lastname: Holland
authors.forename: Maria L
authors.initials: ML
authors.lastname: Torgersen
authors.forename: Kirsten
authors.initials: K
authors.lastname: Sandvig
authors.forename: Anne
authors.initials: A
authors.lastname: Simonsen
mesh_terms: D033963:Adaptor Protein Complex 3 / Q000378:metabolism; D001343:Autophagy; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D002609:Chediak-Higashi Syndrome / Q000378:metabolism*; D004705:Endocytosis; D066246:ErbB Receptors / Q000378:metabolism; D006801:Humans; D008247:Lysosomes / Q000378:metabolism*; D021381:Protein Transport; D059748:Proteolysis; D017527:Receptor, IGF Type 2 / Q000378:metabolism; D033921:Vesicular Transport Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D033963:Adaptor Protein Complex 3; C102161:LYST protein, human; D017527:Receptor, IGF Type 2; D033921:Vesicular Transport Proteins; D066246:ErbB Receptors
keywords: AP-3; CHS; CI-MPR; CtxB; EGFR; LC3; LLPD; LYST; autophagy; beige; cathepsin B; endocytosis; lysosome; ricin; transferrin