The 11S Proteasome Pathway Dependent Regulation Of Tau And Its Function To Alzheimer’s Disease

The REGγ-20S proteasome machinery(also named as 11S proteasome pathway)is a degradation system independent on ubiquitin and ATP,targeting selective substrates,which is effective for tumor,aging,schizophrenia and autoimmune diseases.Initially,we used monoclonal antibody microarray to profile the differential expressed proteins between REGγ knockout and wild-type(WT)mouse embryonic fibroblast(MEF)cells.Significantly,the tau protein level is higher in the REGγknockdown and knockout group,including cell lines(sh-R SH-SY5Y,si-REGγ HT22 and REGγ knockout neuron)and brain tissues(REGγ knockout mice brains).Among them,the sh-R SH-SY5 Y cell line can stably knockdown expressed REGγ created before by transfecting REGγ shRNA and screening from single cell;si-REGγ HT22 line was transfected REGγ silence RNA over 24 hours to knockdown REGγ;REGγknockout neuron was derived from the REGγ knockout mice brain.In addition,it has been proved that REGγ can promote the degradation tau protein in vivo and in vitro,and affect its stability in many kinds of nerve cells by protein degradation assay in vitro and time-course cycloheximide(Chx)assay in vivo.What’s more,the co-immunoprecipitation assay results demonstrated that there was interaction between REGγ and tau protein.It has been previously reported that the mice ablated REGγ had premature aging.Here,we performed western blotting REGγin aging mice hippocampus and PS19 mice hippocampus compared with young and WT group.The AD patients’ hippocampus were also immunohistochemisty stained REGγ compared with the normal healthy human hippocampus.These results indicate that aging associated reduction in REGγ predisposes tauopathy(including neurodegenerative disease like AD).Moreover,the REGγ proteasome mediates direct degradation of both human and mouse tau,predominantly targeting phosphorylated tau proteins(p-tau)that shuttles between cytoplasm and nuclei.REGγ also promotes degradation of tau oligomers that are the more toxic species to tau-related neurodegenerative disease.Furthermore,in order to explore the changes of tau level in the animal model,it has been validated that REGγ-mediated proteasomal degradation of tau in the brain of3～12-month-old REGγ-deficient(REGγ KO),REGγ KO;PS19,and compound mutant mice with conditional neuron specific overexpression in forebrain of REGγ(REGγOE and REGγ OE;PS19)and in PS19.Numerous experiment methods such as nissl staining,western blotting,immunohistochemisty staining,immunoflurorescence staining and golgi staining were performed to show that coupled with tau accumulation(NFT comprised of p-tau)in REGγ-deficiency,neuron loss,dendrite reduction,tau filament accumulation,and microglial activation are much more prominent in the REG γ KO;PS19 than the PS19 model.However,the neuron degenerative lesions are significantly alleviated in the REGγ OE;PS19 mouse model.And then,memory and other behavior analysis(Open field test,Morris Water Maze test,Eight-arm Radial Maze test,Contextual & Cue fear conditioning test,Novel Object Recognition,elevated plus maze and Three-chamber social-approach test)substantiate the role of REGγ in prevention of tauopathy including AD-like symptoms.The cognitive memory impairment could be alleviated in REGγ OE;PS19mouse model compared with REGγ KO;PS19 and PS19 models.The fear and social behaviors were normal in every group of these mouse models from the results of the test of fear conditioning and Three-chamber social-approach.The anxiety level increased in REGγ OE;PS19 mouse model compared with REGγ KO;PS19 and PS19 models showing from the elevated plus maze test.The innovation of this study is that a new mouse model(REGγ OE;PS19 mouse model)was bred,and REGγ decreased in the hippocampus of aging mice,AD mice(PS19 mice)and AD patients,the potential mechanism,TGFβ signaling,associated with REGγ reduction in aging and neuron degeneration conditions was explored.Our study provides valuable insights to new regulatory mechanisms and potentially therapeutic targets for tauopathy and related neurodegenerative diseases represented by tau-induced AD.