14-3-3 binding to LRRK2 is disrupted by multiple Parkinson's disease-associated mutations and regulates cytoplasmic localization.
LRRK2 (leucine-rich repeat protein kinase 2) is mutated in a significant number of Parkinson's disease patients, but still little is understood about how it is regulated or functions. In the present study we have demonstrated that 14-3-3 protein isoforms interact with LRRK2. Consistent with this, endogenous LRRK2 isolated from Swiss ... 3T3 cells or various mouse tissues is associated with endogenous 14-3-3 isoforms. We have established that 14-3-3 binding is mediated by phosphorylation of LRRK2 at two conserved residues (Ser910 and Ser935) located before the leucine-rich repeat domain. Our results suggests that mutation of Ser910 and/or Ser935 to disrupt 14-3-3 binding does not affect intrinsic protein kinase activity, but induces LRRK2 to accumulate within discrete cytoplasmic pools, perhaps resembling inclusion bodies. To investigate links between 14-3-3 binding and Parkinson's disease, we studied how 41 reported mutations of LRRK2 affected 14-3-3 binding and cellular localization. Strikingly, we found that five of the six most common pathogenic mutations (R1441C, R1441G, R1441H, Y1699C and I2020T) display markedly reduced phosphorylation of Ser910/Ser935 thereby disrupting interaction with 14-3-3. We have also demonstrated that Ser910/Ser935 phosphorylation and 14-3-3 binding to endogenous LRRK2 is significantly reduced in tissues of homozygous LRRK2(R1441C) knock-in mice. Consistent with 14-3-3 regulating localization, all of the common pathogenic mutations displaying reduced 14-3-3-binding accumulated within inclusion bodies. We also found that three of the 41 LRRK2 mutations analysed displayed elevated protein kinase activity (R1728H, ~2-fold; G2019S, ~3-fold; and T2031S, ~4-fold). These results provide the first evidence suggesting that 14-3-3 regulates LRRK2 and that disruption of the interaction of LRRK2 with 14-3-3 may be linked to Parkinson's disease.
Mesh Terms:
14-3-3 Proteins, Amino Acid Sequence, Animals, Binding Sites, Brain, Cell Line, Cytoplasm, Green Fluorescent Proteins, Humans, Immunoprecipitation, Kidney, Mice, Mice, Knockout, Microscopy, Fluorescence, Molecular Sequence Data, Mutation, Parkinson Disease, Phosphorylation, Protein Binding, Protein-Serine-Threonine Kinases, Sequence Homology, Amino Acid, Serine, Spleen, Swiss 3T3 Cells
14-3-3 Proteins, Amino Acid Sequence, Animals, Binding Sites, Brain, Cell Line, Cytoplasm, Green Fluorescent Proteins, Humans, Immunoprecipitation, Kidney, Mice, Mice, Knockout, Microscopy, Fluorescence, Molecular Sequence Data, Mutation, Parkinson Disease, Phosphorylation, Protein Binding, Protein-Serine-Threonine Kinases, Sequence Homology, Amino Acid, Serine, Spleen, Swiss 3T3 Cells
Biochem. J.
Date: Sep. 15, 2010
PubMed ID: 20642453
View in: Pubmed Google Scholar
Download Curated Data For This Publication
146342
Switch View:
- Interactions 24