Familial Danish dementia young Knock-in rats expressing humanized APP and human Aβ show impaired pre and postsynaptic glutamatergic transmission

Familial British and Danish dementia (FBD and FDD) are two neurodegenerative disorders caused by mutations in the Integral membrane protein 2B (ITM2b). BRI2, the protein encoded by ITM2b, tunes excitatory synaptic transmission at both pre- and post-synaptic terminus. Too, BRI2 interacts with and modulates proteolytic processing of Amyloid-β precursor Protein (APP), whose mutations cause familial forms of Alzheimer disease (FAD). To study pathogenic mechanism triggered by the Danish mutation we generated rats carrying the Danish mutation into the rat Itm2b gene (Itm2bD rats). Given the BRI2/APP interaction and the widely accepted relevance of human Aβ, a proteolytic product of APP, to AD, Itm2bD rats were engineered to express two humanized App alleles, to produce human Aβ. Here, we studied young Itm2bD rats to investigate early pathogenic changes. We found that peri-adolescent Itm2bD rats present subtle changes in human Aβ levels along with decreased spontaneous glutamate release and AMPAR-mediated responses, but increased short-term synaptic facilitation in the hippocampal Schaeffer-collateral pathway. These changes are similar to those observed in adult mice producing rodent Aβ and carrying either the Danish or British mutations into the mouse Itm2b gene. Collectively, the data show that the pathogenic Danish mutation alters the physiological function of BRI2 at glutamatergic synapses; these functional alterations are detected across species and occur early in life. Future studies will be needed to determine whether this phenomenon represents an early pathogenic event in human dementia.

Structure-based Classification of Tauopathies

Ordered assembly of the tau protein into filaments characterizes multiple neurodegenerative diseases, which are called tauopathies. We previously reported that by electron cryo-microscopy (cryo-EM), tau filament structures from Alzheimer's disease, chronic traumatic encephalopathy (CTE), Pick's disease and corticobasal degeneration (CBD) are distinct. Here we show that the structures of tau filaments from typical and atypical progressive supranuclear palsy (PSP), the most common tauopathy after Alzheimer's disease, define a previously unknown, three-layered fold. Moreover, the tau filament structures from globular glial tauopathy (GGT, Types I and II) are similar to those from PSP. The tau filament fold of argyrophilic grain disease (AGD) differs from the above and resembles the four-layered CBD fold. The majority of tau filaments from aging-related tau astrogliopathy (ARTAG) also have the AGD fold. Surprisingly, tau protofilament structures from inherited cases with mutations +3/+16 in intron 10 of MAPT, the microtubule-associated protein tau gene, are identical to those from AGD, suggesting that a relative overproduction of four-repeat tau can give rise to the AGD fold. Finally, tau filament structures from cases of familial British dementia (FBD) and familial Danish dementia (FDD) are the same as those from Alzheimer's disease and primary age-related tauopathy (PART). These structures provide the basis for a classification of tauopathies that also allows identification of new entities, as we show here for a case diagnosed as PSP, but with abundant spherical 4R tau inclusions in limbic and other brain areas. The structures of the tau fold of this new disease (Limbic-predominant Neuronal inclusion body 4R Tauopathy, LNT) were intermediate between those of GGT and PSP.

A Novel ITM2B Mutation Associated with Familial Chinese Dementia

Mutations in ITM2B have been found to be associated with familial Danish dementia (FDD) and familial British dementia (FBD). Here, we describe a patient with dementia caused by a novel ITM2B p. *267Leuext *11 mutation. The patient presented with dementia, ataxia, deafness, and paraplegia. Amyloid PET and Tau PET showed abnormal deposition of amyloid and tau protein in brain. Summarized from previous 26 FBD and FDD cases, the clinical phenotype of ITM2B; p. *267Leuext *11 mutation in ITM2B is different from the features of FBD and FDD. Our findings increased genetic knowledge of familial dementia and extend the ethnic distribution of ITM2B mutations.

Amyloid and Tau in Neurodegeneration

Background and Hypothesis:Familial British dementia (FBD) and familial Danish dementia (FDD) are two autosomal dominant neurodegenerative diseases caused by mutations in the BRI2 gene. FBD and FDD are characterized by widespread cerebral amyloid angiopathy (CAA), parenchymal amyloid deposition, and neurofibrillary tangles.Previous studies have shown that accumulation of Danish amyloid peptide (ADan) induces hyperphosphorylation of tau and cell cytotoxicity; however, progress in our understanding of the interaction between ADan and tau has been hindered by the lack of antibodies against ADan. In addition, whether the British amyloid peptide (ABri) has a similar effect on tau phosphorylation remains unknown. The main goals of our project are to generate monoclonal antibodies against ADan and to test whether the ABri peptide induces hyperphosphorylation of tau and cell cytotoxicity. Experimental Design or Project Methods:A peptide homologous to the C-terminus of the ADan peptide was used to immunize 5 mice. Serum samples were tested from mice with high ELISA titers and the best 2 animals were used for cell fusion. Cell culture supernatants were screened by ELISA, western blot and immunohistochemistry. ABri peptides were used as negative controls.HEK cells expressing human tau were used to assess the interaction between ABri and tau. We performed immunocytochemistry and Western blot analysis to investigate tau phosphorylation. Results:A total of 11 clones tested positive by western blot. 3 clones tested positive for ABri and were discarded. 5 clones were tested by immunohistochemistry. 3 of the positive clones will be used for subcloning to complete clonality. Work in progress on the interaction between ABri and tau will determine whether ABri induces tau hyperphosphorylation in cell culture. Conclusion and Potential Impact:Our work will provide novel tools to study the interaction between amyloid and tau in neurodegenerative diseases and may uncover possible new targets for pharmaceutical intervention in tauopathies.

Proteomic Characterization of a Mouse Model of Familial Danish Dementia

A dominant mutation in theITM2B/BRI2gene causes familial Danish dementia (FDD) in humans. To model FDD in animal systems, a knock-in approach was recently implemented in mice expressing a wild-type and mutant allele, which bears the FDD-associated mutation. Since theseFDDKImice show behavioural alterations and impaired synaptic function, we characterized their synaptosomal proteome via two-dimensional differential in-gel electrophoresis. After identification by nanoliquid chromatography coupled to electrospray-linear ion trap tandem mass spectrometry, the differentially expressed proteins were classified according to their gene ontology descriptions and their predicted functional interactions. The Dlg4/Psd95 scaffold protein and additional signalling proteins, including protein phosphatases, were revealed by STRING analysis as potential players in the altered synaptic function ofFDDKImice. Immunoblotting analysis finally demonstrated the actual downregulation of the synaptosomal scaffold protein Dlg4/Psd95 and of the dual-specificity phosphatase Dusp3 in the synaptosomes ofFDDKImice.