Aquaporin 4 is differentially increased and dislocated in association with tau and amyloid-beta

AIMS Neurovascular-glymphatic dysfunction plays an important role in Alzheimer's disease and has been analysed mainly in relation to amyloid-beta (Aβ) pathology. Here, we aim to investigate the neurovascular alterations and mapping of aquaporin 4 (AQP4) distribution and dislocation associated with tau and Aβ. MATERIALS AND METHODS Perfusion, susceptibility weighted imaging and structural magnetic resonance imaging (MRI) were performed in the pR5 mouse model of 4-repeat tau and the arcAβ mouse model of amyloidosis. Immunofluorescence staining was performed using antibodies against AQP4, vessel, astroglia, microglia, phospho-tau and Aβ in brain tissue slices from pR5, arcAβ and non-transgenic mice. KEY FINDINGS pR5 mice showed regional atrophy, preserved cerebral blood flow, and reduced cerebral vessel density compared to non-transgenic mice, while arcAβ mice showed cerebral microbleeds and reduced cerebral vessel density. AQP4 dislocation and peri-tau enrichment in the hippocampus and increased AQP4 levels in the cortex and hippocampus were detected in pR5 mice compared to non-transgenic mice. In comparison, cortical AQP4 dislocation and cortical/hippocampal peri-plaque increases were observed in arcAβ mice. Increased expression of reactive astrocytes were detected around the tau inclusions in pR5 mice and Aβ plaques in arcAβ mice. SIGNIFICANCE We demonstrated the neurovascular alterations, microgliosis, astrogliosis and increased AQP4 regional expression in pR5 tau and arcAβ mice. We observed a divergent region-specific AQP4 dislocation and association with phospho-tau and Aβ pathologies

Relationship Between Reactive Astrocytes, by [18F]SMBT-1 Imaging, with Amyloid-Beta, Tau, Glucose Metabolism, and TSPO in Mouse Models of Alzheimer’s Disease

Reactive astrocytes play an important role in the development of Alzheimer’s disease (AD). Here, we aimed to investigate the temporospatial relationships among monoamine oxidase-B, tau and amyloid-β (Aβ), translocator protein, and glucose metabolism by using multitracer imaging in AD transgenic mouse models. Positron emission tomography (PET) imaging with [18F]SMBT-1 (monoamine oxidase-B), [18F]florbetapir (Aβ), [18F]PM-PBB3 (tau), [18F]fluorodeoxyglucose (FDG), and [18F]DPA-714 (translocator protein) was carried out in 5- and 10-month-old APP/PS1, 11-month-old 3×Tg mice, and aged-matched wild-type mice. The brain regional referenced standard uptake value (SUVR) was computed with the cerebellum as the reference region. Immunofluorescence staining was performed on mouse brain tissue slices. [18F]SMBT-1 and [18F]florbetapir SUVRs were greater in the cortex and hippocampus of 10-month-old APP/PS1 mice than in those of 5-month-old APP/PS1 mice and wild-type mice. No significant difference in the regional [18F]FDG or [18F]DPA-714 SUVRs was observed in the brains of 5- or 10-month-old APP/PS1 mice or wild-type mice. No significant difference in the SUVRs of any tracer was observed between 11-month-old 3×Tg mice and age-matched wild-type mice. A positive correlation between the SUVRs of [18F]florbetapir and [18F]DPA-714 in the cortex and hippocampus was observed among the transgenic mice. Immunostaining validated the distribution of MAO-B and limited Aβ and tau pathology in 11-month-old 3×Tg mice; and Aβ deposits in brain tissue from 10-month-old APP/PS1 mice. In summary, these findings provide in vivo evidence that an increase in astrocyte [18F]SMBT-1 accompanies Aβ accumulation in APP/PS1 models of AD amyloidosis

Gains and losses on the road to understanding Alzheimer's disease

Alzheimer's disease (AD) is a neurodegenerative disorder and the most common cause for dementia, which affects approximately 120 thousand people in Switzerland and 35 million worldwide. Aging is a major risk factor for developing AD and thus, as our societies are growing older, we face great challenges to find treatment strategies. The disease is characterised by loss of memory, deposition of extracellular amyloid plaques containing Aβ peptides and intraneuronal tangles of the tau protein. To date, there is no effective treatment and the cause of the disease is still debated. The Schweizerische Alzheimervereinigung states that we need "continuous manifold research" into all possible causes of AD to find a cure for this disease. Fitting this proposition, a recent publication by Xia et al. (2015) described a novel mouse model that for the first time reproduces cortical neuron death as observed in human AD cases. At the same time, this publication questions the major theory of AD pathogenesis and points towards different treatment avenues that should be followed to find a cure for AD

Fe65 is the sole member of its family that mediates transcription regulated by the amyloid precursor protein

The amyloid precursor protein (APP), a central molecule in Alzheimer's disease (AD), has physiological roles in cell adhesion and signaling, migration, neurite outgrowth and synaptogenesis. Intracellular adapter proteins mediate the function of transmembrane proteins. Fe65 (also known as APBB1) is a major APP-binding protein. Regulated intramembrane proteolysis (RIP) by γ-secretase releases the APP intracellular domain (AICD), together with the interacting proteins, from the membrane. We studied the impact of the Fe65 family (Fe65, and its homologs Fe65L1 and Fe65L2, also known as APBB2 and APBB3, respectively) on the nuclear signaling function of the AICD. All Fe65 family members increased amyloidogenic processing of APP, generating higher levels of β-cleaved APP stubs and AICD. However, Fe65 was the only family member supporting AICD translocation to nuclear spots and its transcriptional activity. Using a recently established transcription assay, we dissected the transcriptional activity of Fe65 and provide strong evidence that Fe65 represents a transcription factor. We show that Fe65 relies on the lysine acetyltransferase Tip60 (also known as KAT5) for nuclear translocation. Furthermore, inhibition of APP cleavage reduces nuclear Tip60 levels, but this does not occur in Fe65-knockout cells. The rate of APP cleavage therefore regulates the nuclear translocation of AICD-Fe65-Tip60 (AFT) complexes, to promote transcription by Fe65

A fluorescent protein-readout for transcriptional activity reveals regulation of APP nuclear signaling by phosphorylation sites

Signaling pathways that originate at the plasma membrane, including regulated intramembrane proteolysis (RIP), enable extracellular cues to control transcription. We modified the yeast Gal4 transcription system to study the nuclear translocation of transcriptionally active complexes using the fluorescent protein citrine (Cit) as a reporter. This enabled highly sensitive quantitative analysis of transcription in situ at the single cell level. The Gal4/UAS-Cit transcription assay displayed a sigmoidal response limited by the number of integrated reporter cassettes. We validated the assay by analyzing nuclear translocation of the amyloid precursor protein (APP) intracellular domain (AICD) and confirmed the requirement of Fe65 for nuclear translocation of AICD. In addition to the strong on-off effects on transcriptional activity, the results of this assay establish that phosphorylation modifies nuclear signaling. The Y682F mutation in APP showed the strongest increase in Cit expression, underscoring its role in regulating Fe65 binding. Together, we established a highly sensitive fluorescent protein-based assay that can monitor transcriptional activity at the single cell level and demonstrate that AICD phosphorylation affects Fe65 nuclear activity. This assay also introduces a platform for future single cell-based drug screening methods for nuclear translocation

Lysine acetyltransferase Tip60 acetylates the APP adaptor Fe65 to increase its transcriptional activity

Proteolytic processing of the amyloid precursor protein (APP) releases the APP intracellular domain (AICD) from the membrane. Bound to the APP adaptor protein Fe65 and the lysine acetyltransferase (KAT) Tip60, AICD translocates to the nucleus. Here, the complex forms spherical condensates at sites of endogenous target genes, termed AFT spots (AICD-Fe65-Tip60). We show that loss of Tip60 KAT activity prevents autoacetylation, reduces binding of Fe65 and abolishes Fe65-mediated stabilization of Tip60. Autoacetylation is a prerequisite for AFT spot formation, with KAT-deficient Tip60 retained together with Fe65 in speckles. We identify lysine residues 204 and 701 of Fe65 as acetylation targets of Tip60. We do not detect acetylation of AICD. Mutation of Fe65 K204 and K701 to glutamine, mimicking acetylation-induced charge neutralization, increases the transcriptional activity of Fe65 whereas Tip60 inhibition reduces it. The lysine deacetylase (KDAC) class III Sirt1 deacetylates Fe65 and pharmacological modulation of Sirt1 activity regulates Fe65 transcriptional activity. A second acetylation/deacetylation cycle, conducted by CBP and class I/II KDACs at different lysine residues, regulates stability of Fe65. This is the first report describing a role for acetylation in the regulation of Fe65 transcriptional activity, with Tip60 being the only KAT tested that supports AFT spot formation

Fe65-PTB2 Dimerization Mimics Fe65-APP Interaction

Physiological function and pathology of the Alzheimer's disease causing amyloid precursor protein (APP) are correlated with its cytosolic adaptor Fe65 encompassing a WW and two phosphotyrosine-binding domains (PTBs). The C-terminal Fe65-PTB2 binds a large portion of the APP intracellular domain (AICD) including the GYENPTY internalization sequence fingerprint. AICD binding to Fe65-PTB2 opens an intra-molecular interaction causing a structural change and altering Fe65 activity. Here we show that in the absence of the AICD, Fe65-PTB2 forms a homodimer in solution and determine its crystal structure at 2.6 Å resolution. Dimerization involves the unwinding of a C-terminal α-helix that mimics binding of the AICD internalization sequence, thus shielding the hydrophobic binding pocket. Specific dimer formation is validated by nuclear magnetic resonance (NMR) techniques and cell-based analyses reveal that Fe65-PTB2 together with the WW domain are necessary and sufficient for dimerization. Together, our data demonstrate that Fe65 dimerizes via its APP interaction site, suggesting that besides intra- also intermolecular interactions between Fe65 molecules contribute to homeostatic regulation of APP mediated signaling

Relationship Between Reactive Astrocytes, by [18F]SMBT-1 Imaging, with Amyloid-Beta, Tau, Glucose Metabolism, and TSPO in Mouse Models of Alzheimer's Disease.

Reactive astrocytes play an important role in the development of Alzheimer's disease (AD). Here, we aimed to investigate the temporospatial relationships among monoamine oxidase-B, tau and amyloid-β (Aβ), translocator protein, and glucose metabolism by using multitracer imaging in AD transgenic mouse models. Positron emission tomography (PET) imaging with [18F]SMBT-1 (monoamine oxidase-B), [18F]florbetapir (Aβ), [18F]PM-PBB3 (tau), [18F]fluorodeoxyglucose (FDG), and [18F]DPA-714 (translocator protein) was carried out in 5- and 10-month-old APP/PS1, 11-month-old 3×Tg mice, and aged-matched wild-type mice. The brain regional referenced standard uptake value (SUVR) was computed with the cerebellum as the reference region. Immunofluorescence staining was performed on mouse brain tissue slices. [18F]SMBT-1 and [18F]florbetapir SUVRs were greater in the cortex and hippocampus of 10-month-old APP/PS1 mice than in those of 5-month-old APP/PS1 mice and wild-type mice. No significant difference in the regional [18F]FDG or [18F]DPA-714 SUVRs was observed in the brains of 5- or 10-month-old APP/PS1 mice or wild-type mice. No significant difference in the SUVRs of any tracer was observed between 11-month-old 3×Tg mice and age-matched wild-type mice. A positive correlation between the SUVRs of [18F]florbetapir and [18F]DPA-714 in the cortex and hippocampus was observed among the transgenic mice. Immunostaining validated the distribution of MAO-B and limited Aβ and tau pathology in 11-month-old 3×Tg mice; and Aβ deposits in brain tissue from 10-month-old APP/PS1 mice. In summary, these findings provide in vivo evidence that an increase in astrocyte [18F]SMBT-1 accompanies Aβ accumulation in APP/PS1 models of AD amyloidosis

Aquaporin 4 is differentially increased and dislocated in association with tau and amyloid-beta

Aims: Neurovascular-glymphatic dysfunction plays an important role in Alzheimer's disease and has been analysed mainly in relation to amyloid-beta (Aβ) pathology. Here, we aim to investigate the neurovascular alterations and mapping of aquaporin 4 (AQP4) distribution and dislocation associated with tau and Aβ. Materials and methods: Perfusion, susceptibility weighted imaging and structural magnetic resonance imaging (MRI) were performed in the pR5 mouse model of 4-repeat tau and the arcAβ mouse model of amyloidosis. Immunofluorescence staining was performed using antibodies against AQP4, vessel, astroglia, microglia, phospho-tau and Aβ in brain tissue slices from pR5, arcAβ and non-transgenic mice. Key findings: pR5 mice showed regional atrophy, preserved cerebral blood flow, and reduced cerebral vessel density compared to non-transgenic mice, while arcAβ mice showed cerebral microbleeds and reduced cerebral vessel density. AQP4 dislocation and peri-tau enrichment in the hippocampus and increased AQP4 levels in the cortex and hippocampus were detected in pR5 mice compared to non-transgenic mice. In comparison, cortical AQP4 dislocation and cortical/hippocampal peri-plaque increases were observed in arcAβ mice. Increased expression of reactive astrocytes were detected around the tau inclusions in pR5 mice and Aβ plaques in arcAβ mice. Significance: We demonstrated the neurovascular alterations, microgliosis, astrogliosis and increased AQP4 regional expression in pR5 tau and arcAβ mice. We observed a divergent region-specific AQP4 dislocation and association with phospho-tau and Aβ pathologies.ISSN:0024-3205ISSN:1879-063

Early alterations in functional connectivity and white matter structure in a transgenic mouse model of cerebral amyloidosis

Impairment of brain functional connectivity (FC) is thought to be an early event occurring in diseases with cerebral amyloidosis, such as Alzheimer's disease. Regions sustaining altered functional networks have been shown to colocalize with regions marked with amyloid plaques burden suggesting a strong link between FC and amyloidosis. Whether the decline in FC precedes amyloid plaque deposition or is a consequence thereof is currently unknown. The sequence of events during early stages of the disease is difficult to capture in humans due to the difficulties in providing an early diagnosis and also in view of the heterogeneity among patients. Transgenic mouse lines overexpressing amyloid precursor proteins develop cerebral amyloidosis and constitute an attractive model system for studying the relationship between plaque and functional changes. In this study, ArcAβ transgenic and wild-type mice were imaged using resting-state fMRI methods across their life-span in a cross-sectional design to analyze changes in FC in relation to the pathology. Transgenic mice show compromised development of FC during the first months of postnatal life compared with wild-type animals, resulting in functional impairments that affect in particular the sensory-motor cortex already in preplaque stage. These functional alterations were accompanied by structural changes as reflected by reduced fractional anisotropy values, as derived from diffusion tensor imaging. Our results suggest cerebral amyloidosis in mice is preceded by impairment of neuronal networks and white matter structures. FC analysis in mice is an attractive tool for studying the implications of impaired neuronal networks in models of cerebral amyloid pathology