title: Reevaluation of phosphorylation sites in the Parkinson disease-associated leucine-rich repeat kinase 2.
abstract: Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene have been identified as an important cause of late-onset, autosomal dominant familial Parkinson disease and contribute to sporadic Parkinson disease. LRRK2 is a large complex protein with multiple functional domains, including a Roc-GTPase, protein kinase, and multiple protein-protein interaction domains. Previous studies have suggested an important role for kinase activity in LRRK2-induced neuronal toxicity and inclusion body formation. Disease-associated mutations in LRRK2 also tend to increase kinase activity. Thus, enhanced kinase activity may therefore underlie LRRK2-linked disease. Similar to the closely related mixed-lineage kinases, LRRK2 can undergo autophosphorylation in vitro. Three putative autophosphorylation sites (Thr-2031, Ser-2032, and Thr-2035) have been identified within the activation segment of the LRRK2 kinase domain based on sequence homology to mixed-lineage kinases. Phosphorylation at one or more of these sites is critical for the kinase activity of LRRK2. Sensitive phospho-specific antibodies to each of these three sites have been developed and validated by ELISA, dot-blot, and Western blot analysis. Using these antibodies, we have found that all three putative sites are phosphorylated in LRRK2, and Ser-2032 and Thr-2035 are the two important sites that regulate LRRK2 kinase activity.
authors.affiliation: NeuroRegeneration and Stem Cell Programs, Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
authors.forename: Xiaojie
authors.initials: X
authors.lastname: Li
authors.forename: Darren J
authors.initials: DJ
authors.lastname: Moore
authors.forename: Yulan
authors.initials: Y
authors.lastname: Xiong
authors.forename: Ted M
authors.initials: TM
authors.lastname: Dawson
authors.forename: Valina L
authors.initials: VL
authors.lastname: Dawson
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D045163:Antibodies, Phospho-Specific / Q000235:genetics / Q000276:immunology; D015153:Blotting, Western; D002460:Cell Line; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D000071158:Leucine-Rich Repeat Serine-Threonine Protein Kinase-2; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D010300:Parkinson Disease / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
chemical_list: D045163:Antibodies, Phospho-Specific; D006861:Hydrogen Peroxide; C495280:LRRK2 protein, human; D000071158:Leucine-Rich Repeat Serine-Threonine Protein Kinase-2; C495279:Lrrk2 protein, mouse; D017346:Protein Serine-Threonine Kinases