Effects of DJ-1 mutations and polymorphisms on protein stability and subcellular localization.
Mutations in the DJ-1 gene are associated with recessive, early onset Parkinson's disease (PD). We reported previously that one of the point mutations, L166P, destabilizes the protein and thus produces an effective knockout of the gene. Here, we have expanded this analysis to include a series of mutations and polymorphisms identified throughout the gene. The M26I point mutation was also unstable, although the effect was not as dramatic as with L166P. Protein levels were rescued in part, but not completely, by proteasome inhibition. Other variants, such as R98Q, were generally stable. We noted that M26I and L166P are both in helical regions near the dimer interface. However, M26I retains the ability to dimerize. We also examined the subcellular localization of DJ-1 and found that most mutations were similar to the wild-type (wt) protein in that a few cells showed mitochondrial staining. However, in all cases, the proportion of cells with mitochondrial DJ-1 staining was increased in oxidative conditions, suggesting that oxidation promotes the mitochondrial localization of DJ-1.
Blackinton
Jeff
J
Laboratory of Neurogenetics, National Institute on Aging, NIH, Building 10 Room 6C103, MSC1589 9000 Rockville Pike, Bethesda, MD 20892, USA.
Ahmad
Rili
R
Miller
David W
DW
van der Brug
Marcel P
MP
Canet-Avilés
Rosa M
RM
Hague
Stephen M
SM
Kaleem
Mona
M
Cookson
Mark R
MR
D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000643:Ammonium Chloride / Q000494:pharmacology; D000704:Analysis of Variance; D000818:Animals; D015153:Blotting, Western / Q000379:methods; D045744:Cell Line, Tumor; D003513:Cycloheximide / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D019281:Dimerization; D005455:Fluorescent Antibody Technique / Q000379:methods; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation / Q000379:methods; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D008958:Models, Molecular; D009447:Neuroblastoma; D015513:Oncogene Proteins / Q000737:chemistry / Q000235:genetics*; D017354:Point Mutation; D011110:Polymorphism, Genetic; D000071617:Protein Deglycase DJ-1; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013347:Subcellular Fractions / Q000378:metabolism; D013997:Time Factors; D014162:Transfection / Q000379:methods
D015853:Cysteine Proteinase Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D015513:Oncogene Proteins; D011500:Protein Synthesis Inhibitors; D012333:RNA, Messenger; D000643:Ammonium Chloride; C067713:lactacystin; D049452:Green Fluorescent Proteins; D003513:Cycloheximide; C105131:PARK7 protein, human; D000071617:Protein Deglycase DJ-1; D000111:Acetylcysteine