title: Chaperone-mediated autophagy is defective in mucolipidosis type IV.
abstract: Mucolipidosis type IV (MLIV) is a lysosomal storage disorder caused by mutations in the MCOLN1 gene, a member of the transient receptor potential (TRP) cation channel gene family. The encoded protein, transient receptor potential mucolipin-1 (TRPML1), has been localized to lysosomes and late endosomes but the pathogenic mechanism by which loss of TRPML1 leads to abnormal cellular storage and neuronal cell death is still poorly understood. Yeast two-hybrid and co-immunoprecipitation (coIP) experiments identified interactions between TRPML1 and Hsc70 as well as TRPML1 and Hsp40. Hsc70 and Hsp40 are members of a molecular chaperone complex required for protein transport into the lysosome during chaperone-mediated autophagy (CMA). To determine the functional relevance of this interaction, we compared fibroblasts from MLIV patients to those from sex- and age-matched controls and show a defect in CMA in response to serum withdrawal. This defect in CMA was subsequently confirmed in purified lysosomes isolated from control and MLIV fibroblasts. We further show that the amount of lysosomal-associated membrane protein type 2A (LAMP-2A) is reduced in lysosomal membranes of MLIV fibroblasts. As a result of decreased CMA, MLIV fibroblasts have increased levels of oxidized proteins compared to control fibroblasts. We hypothesize that TRPML1 may act as a docking site for intralysosomal Hsc70 (ly-Hsc70) allowing it to more efficiently pull in substrates for CMA. It is also possible that TRPML1 channel activity may be required for CMA. Understanding the role of TRPML1 in CMA will undoubtedly help to characterize the pathogenesis of MLIV.
authors.affiliation: Center for Human Genetic Research, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts 02114, USA.
authors.forename: Bhuvarahamurthy
authors.initials: B
authors.lastname: Venugopal
authors.forename: Nicholas T
authors.initials: NT
authors.lastname: Mesires
authors.forename: John C
authors.initials: JC
authors.lastname: Kennedy
authors.forename: Cyntia
authors.initials: C
authors.lastname: Curcio-Morelli
authors.forename: Janice M
authors.initials: JM
authors.lastname: Laplante
authors.forename: J Fred
authors.initials: JF
authors.lastname: Dice
authors.forename: Susan A
authors.initials: SA
authors.lastname: Slaugenhaupt
mesh_terms: D000818:Animals; D001343:Autophagy / Q000502:physiology*; D016466:CHO Cells; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D005347:Fibroblasts / Q000166:cytology / Q000502:physiology; D050883:HSC70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D050956:HSP40 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D018841:HSP90 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D015759:Ionomycin / Q000378:metabolism; D007476:Ionophores / Q000378:metabolism; D052119:Lysosomal-Associated Membrane Protein 2; D051907:Lysosomal Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008247:Lysosomes / Q000378:metabolism / Q000648:ultrastructure; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism*; D009081:Mucolipidoses / Q000235:genetics / Q000378:metabolism* / Q000503:physiopathology*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D050053:TRPM Cation Channels / Q000235:genetics / Q000378:metabolism*; D050051:Transient Receptor Potential Channels / Q000235:genetics / Q000378:metabolism; D020798:Two-Hybrid System Techniques
chemical_list: D050883:HSC70 Heat-Shock Proteins; D050956:HSP40 Heat-Shock Proteins; D018841:HSP90 Heat-Shock Proteins; D007476:Ionophores; C497811:LAMP2 protein, human; D052119:Lysosomal-Associated Membrane Protein 2; D051907:Lysosomal Membrane Proteins; C416674:MCOLN1 protein, human; C510642:MCOLN3 protein, human; C510641:Mcoln2 protein, human; D018832:Molecular Chaperones; D011993:Recombinant Fusion Proteins; D050053:TRPM Cation Channels; D050051:Transient Receptor Potential Channels; D015759:Ionomycin