title: Glycosylation, transport, and complex formation of palmitoyl protein thioesterase 1 (PPT1)--distinct characteristics in neurons.
abstract: BACKGROUND
  Neuronal ceroid lipofuscinoses (NCLs) are collectively the most common type of recessively inherited childhood encephalopathies. The most severe form of NCL, infantile neuronal ceroid lipofuscinosis (INCL), is caused by mutations in the CLN1 gene, resulting in a deficiency of the lysosomal enzyme, palmitoyl protein thioesterase 1 (PPT1). The deficiency of PPT1 causes a specific death of neocortical neurons by a mechanism, which is currently unclear. To understand the function of PPT1 in more detail, we have further analyzed the basic properties of the protein, especially focusing on possible differences in non-neuronal and neuronal cells.
  
  
  RESULTS
  Our study shows that the N-glycosylation of N197 and N232, but not N212, is essential for PPT1's activity and intracellular transport. Deglycosylation of overexpressed PPT1 produced in neurons and fibroblasts demonstrates differentially modified PPT1 in different cell types. Furthermore, antibody internalization assays showed differences in PPT1 transport when compared with a thoroughly characterized lysosomal enzyme aspartylglucosaminidase (AGA), an important observation potentially influencing therapeutic strategies. PPT1 was also demonstrated to form oligomers by size-exclusion chromatography and co-immunoprecipitation assays. Finally, the consequences of disease mutations were analyzed in the perspective of our new results, suggesting that the mutations increase both the degree of glycosylation of PPT1 and its ability to form complexes.
  
  
  CONCLUSIONS
  Our current study describes novel properties for PPT1. We observe differences in PPT1 processing and trafficking in neuronal and non-neuronal cells, and describe for the first time the ability of PPT1 to form complexes. Understanding the basic characteristics of PPT1 is fundamental in order to clarify the molecular pathogenesis behind neurodegeneration in INCL.
authors.affiliation: Department of Molecular Medicine, National Public Health Institute, Biomedicum Helsinki, Haartmaninkatu 8, Helsinki, Finland. annina.lyly@ktl.fi <annina.lyly@ktl.fi>
authors.forename: Annina
authors.initials: A
authors.lastname: Lyly
authors.forename: Carina
authors.initials: C
authors.lastname: von Schantz
authors.forename: Tarja
authors.initials: T
authors.lastname: Salonen
authors.forename: Outi
authors.initials: O
authors.lastname: Kopra
authors.forename: Jani
authors.initials: J
authors.lastname: Saarela
authors.forename: Matti
authors.initials: M
authors.lastname: Jauhiainen
authors.forename: Aija
authors.initials: A
authors.lastname: Kyttälä
authors.forename: Anu
authors.initials: A
authors.lastname: Jalanko
mesh_terms: D000818:Animals; D001227:Aspartylglucosylaminase / Q000378:metabolism; D019556:COS Cells; D018929:Cell Culture Techniques; D002522:Chlorocebus aethiops; D006031:Glycosylation; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D009154:Mutation; D009474:Neurons / Q000166:cytology / Q000201:enzymology / Q000502:physiology*; D009928:Organ Specificity; D016716:PC12 Cells; D021381:Protein Transport; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D013869:Thiolester Hydrolases / Q000378:metabolism*
chemical_list: D008565:Membrane Proteins; D011994:Recombinant Proteins; D013869:Thiolester Hydrolases; C416054:PPT1 protein, human; C095299:palmitoyl-protein thioesterase; D001227:Aspartylglucosylaminase