title: Macroautophagy is defective in mucolipin-1-deficient mouse neurons.
abstract: Mucolipidosis type IV is a neurodegenerative lysosomal disease clinically characterized by psychomotor retardation, visual impairment, and achlorhydria. In this study we report the development of a neuronal cell model generated from cerebrum of Mcoln1(-/-) embryos. Prior functional characterization of MLIV cells has been limited to fibroblast cultures gleaned from patients. The current availability of the mucolipin-1 knockout mouse model Mcoln1(-/-) allows the study of mucolipin-1-defective neurons, which is important since the disease is characterized by severe neurological impairment. Electron microscopy studies reveal significant membranous intracytoplasmic storage bodies, which correlate with the storage morphology observed in cerebral cortex of Mcoln1(-/-) P7 pups and E17 embryos. The Mcoln1(-/-) neuronal cultures show an increase in size of LysoTracker and Lamp1 positive vesicles. Using this neuronal model system, we show that macroautophagy is defective in mucolipin-1-deficient neurons and that LC3-II levels are significantly elevated. Treatment with rapamycin plus protease inhibitors did not increase levels of LC3-II in Mcoln1(-/-) neuronal cultures, indicating that the lack of mucolipin-1 affects LC3-II clearance. P62/SQSTM1 and ubiquitin levels were also increased in Mcoln1(-/-) neuronal cultures, suggesting an accumulation of protein aggregates and a defect in macroautophagy which could help explain the neurodegeneration observed in MLIV. This study describes, for the first time, a defect in macroautophagy in mucolipin-1-deficient neurons, which corroborates recent findings in MLIV fibroblasts and provides new insight into the neuronal pathogenesis of this disease.
authors.affiliation: Center for Human Genetic Research, Massachusetts General Hospital/Harvard Medical School, Boston, MA 02114, USA.
authors.forename: Cyntia
authors.initials: C
authors.lastname: Curcio-Morelli
authors.forename: Florie A
authors.initials: FA
authors.lastname: Charles
authors.forename: Matthew C
authors.initials: MC
authors.lastname: Micsenyi
authors.forename: Yi
authors.initials: Y
authors.lastname: Cao
authors.forename: Bhuvarahamurthy
authors.initials: B
authors.lastname: Venugopal
authors.forename: Marsha F
authors.initials: MF
authors.lastname: Browning
authors.forename: Kostantin
authors.initials: K
authors.lastname: Dobrenis
authors.forename: Susan L
authors.initials: SL
authors.lastname: Cotman
authors.forename: Steven U
authors.initials: SU
authors.lastname: Walkley
authors.forename: Susan A
authors.initials: SA
authors.lastname: Slaugenhaupt
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000588:Amines / Q000378:metabolism; D000818:Animals; D001343:Autophagy; D002478:Cells, Cultured; D006360:Heat-Shock Proteins / Q000378:metabolism; D051907:Lysosomal Membrane Proteins / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D009081:Mucolipidoses / Q000235:genetics / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism* / Q000473:pathology* / Q000648:ultrastructure; D000071456:Sequestosome-1 Protein; D050053:TRPM Cation Channels / Q000172:deficiency* / Q000235:genetics; D051758:Transcription Factor TFIIH; D014157:Transcription Factors / Q000378:metabolism; D050051:Transient Receptor Potential Channels; D025801:Ubiquitin / Q000378:metabolism; D052118:Lysosomal-Associated Membrane Protein 1
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D000588:Amines; D006360:Heat-Shock Proteins; D051907:Lysosomal Membrane Proteins; D008869:Microtubule-Associated Proteins; D000071456:Sequestosome-1 Protein; D050053:TRPM Cation Channels; D051758:Transcription Factor TFIIH; D014157:Transcription Factors; D050051:Transient Receptor Potential Channels; D025801:Ubiquitin; C490981:Gtf2h1 protein, mouse; C052255:Lamp1 protein, mouse; D052118:Lysosomal-Associated Membrane Protein 1; C480860:Map1lc3b protein, mouse; C512321:Mcoln1 protein, mouse; C493330:Red DND-99; C101343:Sqstm1 protein, mouse