Mikhail A. Kostylev

Alzheimer amyloid-β oligomer bound to postsynaptic prion protein activates Fyn to impair neurons.

Amyloid-beta (Aβ) oligomers are thought to trigger Alzheimers disease pathophysiology. Cellular prion protein (PrPC) selectively binds oligomeric Aβ and can mediate Alzheimers disease–related phenotypes. We examined the specificity, distribution and signaling of Aβ-PrPC complexes, seeking to understand how they might alter the function of NMDA receptors (NMDARs) in neurons. PrPC is enriched in postsynaptic densities, and Aβ-PrPC interaction leads to Fyn kinase activation. Soluble Aβ assemblies derived from the brains of individuals with Alzheimers disease interacted with PrPC to activate Fyn. Aβ engagement of PrPC-Fyn signaling yielded phosphorylation of the NR2B subunit of NMDARs, which was coupled to an initial increase and then a loss of surface NMDARs. Aβ-induced dendritic spine loss and lactate dehydrogenase release required both PrPC and Fyn, and human familial Alzheimers disease transgene–induced convulsive seizures did not occur in mice lacking PrPC. These results delineate an Aβ oligomer signal transduction pathway that requires PrPC and Fyn to alter synaptic function, with deleterious consequences in Alzheimers disease.

Metabotropic Glutamate Receptor 5 Is a Coreceptor for Alzheimer Aβ Oligomer Bound to Cellular Prion Protein

Soluble amyloid-β oligomers (Aβo) trigger Alzheimers disease (AD) pathophysiology and bind with high affinity to cellular prion protein (PrP(C)). At the postsynaptic density (PSD), extracellular Aβo bound to lipid-anchored PrP(C) activates intracellular Fyn kinase to disrupt synapses. Here, we screened transmembrane PSD proteins heterologously for the ability to couple Aβo-PrP(C) with Fyn. Only coexpression of the metabotropic glutamate receptor, mGluR5, allowed PrP(C)-bound Aβo to activate Fyn. PrP(C) and mGluR5 interact physically, and cytoplasmic Fyn forms a complex with mGluR5. Aβo-PrP(C) generates mGluR5-mediated increases of intracellular calcium in Xenopus oocytes and in neurons, and the latter is also driven by human AD brain extracts. In addition, signaling by Aβo-PrP(C)-mGluR5 complexes mediates eEF2 phosphorylation and dendritic spine loss. For mice expressing familial AD transgenes, mGluR5 antagonism reverses deficits in learning, memory, and synapse density. Thus, Aβo-PrP(C) complexes at the neuronal surface activate mGluR5 to disrupt neuronal function.

Fyn inhibition rescues established memory and synapse loss in Alzheimer mice

Currently no effective disease‐modifying agents exist for the treatment of Alzheimer disease (AD). The Fyn tyrosine kinase is implicated in AD pathology triggered by amyloid‐ß oligomers (Aßo) and propagated by Tau. Thus, Fyn inhibition may prevent or delay disease progression. Here, we sought to repurpose the Src family kinase inhibitor oncology compound, AZD0530, for AD.

Prion-Protein-interacting Amyloid-β Oligomers of High Molecular Weight Are Tightly Correlated with Memory Impairment in Multiple Alzheimer Mouse Models

Background: Amyloid-β (Aβ) oligomers are key in Alzheimer disease (AD) but are diverse and poorly characterized. Results: Multiple Aβ forms were measured across the life span of AD model mice and human AD brain. Conclusion: Aβ species interacting with prion protein were tightly linked to behavioral impairment. Significance: An Aβ oligomer subset with defined biochemical properties is present in multiple AD-relevant samples. Alzheimer disease (AD) is characterized by amyloid-β accumulation, with soluble oligomers (Aβo) being the most synaptotoxic. However, the multivalent and unstable nature of Aβo limits molecular characterization and hinders research reproducibility. Here, we characterized multiple Aβo forms throughout the life span of various AD mice and in post-mortem human brain. Aβo exists in several populations, where prion protein (PrPC)-interacting Aβo is a high molecular weight Aβ assembly present in multiple mice and humans with AD. Levels of PrPC-interacting Aβo match closely with mouse memory and are equal or superior to other Aβ measures in predicting behavioral impairment. However, Aβo metrics vary considerably between mouse strains. Deleting PrPC expression in mice with relatively low PrPC-interacting Aβo (Tg2576) results in partial rescue of cognitive performance as opposed to complete recovery in animals with a high percentage of PrPC-interacting Aβo (APP/PSEN1). These findings highlight the relative contributions and interplay of Aβo forms in AD.

Metabotropic glutamate receptor 5 couples cellular prion protein to intracellular signalling in Alzheimer’s disease

Alzheimers disease-related phenotypes in mice can be rescued by blockade of either cellular prion protein or metabotropic glutamate receptor 5. We sought genetic and biochemical evidence that these proteins function cooperatively as an obligate complex in the brain. We show that cellular prion protein associates via transmembrane metabotropic glutamate receptor 5 with the intracellular protein mediators Homer1b/c, calcium/calmodulin-dependent protein kinase II, and the Alzheimers disease risk gene product protein tyrosine kinase 2 beta. Coupling of cellular prion protein to these intracellular proteins is modified by soluble amyloid-β oligomers, by mouse brain Alzheimers disease transgenes or by human Alzheimers disease pathology. Amyloid-β oligomer-triggered phosphorylation of intracellular protein mediators and impairment of synaptic plasticity in vitro requires Prnp-Grm5 genetic interaction, being absent in transheterozygous loss-of-function, but present in either single heterozygote. Importantly, genetic coupling between Prnp and Grm5 is also responsible for signalling, for survival and for synapse loss in Alzheimers disease transgenic model mice. Thus, the interaction between metabotropic glutamate receptor 5 and cellular prion protein has a central role in Alzheimers disease pathogenesis, and the complex is a potential target for disease-modifying intervention.

Therapeutic molecules and endogenous ligands regulate the interaction between brain cellular prion protein (PrPC) and metabotropic glutamate receptor 5 (mGluR5)

Background: Amyloid-β oligomers trigger Alzheimer disease pathophysiology via the interaction of cellular prion protein (PrPC) with metabotropic glutamate receptor 5 (mGluR5). Results: PrPC region 91–153 interacts preferentially with the activated conformation of mGluR5. Conclusion: Antibodies against PrPC region 91–153 and agonist/antagonist-driven mGluR5 conformations regulate the PrPC-mGluR5 interaction. Significance: These findings have therapeutic implications for Alzheimer disease by identifying compounds that modulate the PrPC-mGluR5 interaction. Soluble Amyloid-β oligomers (Aβo) can trigger Alzheimer disease (AD) pathophysiology by binding to cell surface cellular prion protein (PrPC). PrPC interacts physically with metabotropic glutamate receptor 5 (mGluR5), and this interaction controls the transmission of neurotoxic signals to intracellular substrates. Because the interruption of the signal transduction from PrPC to mGluR5 has therapeutic potential for AD, we developed assays to explore the effect of endogenous ligands, agonists/antagonists, and antibodies on the interaction between PrPC and mGluR5 in cell lines and mouse brain. We show that the PrPC segment of amino acids 91–153 mediates the interaction with mGluR5. Agonists of mGluR5 increase the mGluR5-PrPC interaction, whereas mGluR5 antagonists suppress protein association. Synthetic Aβo promotes the protein interaction in mouse brain and transfected HEK-293 cell membrane preparations. The interaction of PrPC and mGluR5 is enhanced dramatically in the brains of familial AD transgenic model mice. In brain homogenates with Aβo, the interaction of PrPC and mGluR5 is reversed by mGluR5-directed antagonists or antibodies directed against the PrPC segment of amino acids 91–153. Silent allosteric modulators of mGluR5 do not alter Glu or basal mGluR5 activity, but they disrupt the Aβo-induced interaction of mGluR5 with PrPC. The assays described here have the potential to identify and develop new compounds that inhibit the interaction of PrPC and mGluR5, which plays a pivotal role in the pathogenesis of Alzheimer disease by transmitting the signal from extracellular Aβo into the cytosol.

Brivaracetam, but not ethosuximide, reverses memory impairments in an Alzheimer's disease mouse model.

IntroductionRecent studies have shown that several strains of transgenic Alzheimer’s disease (AD) mice overexpressing the amyloid precursor protein (APP) have cortical hyperexcitability, and their results have suggested that this aberrant network activity may be a mechanism by which amyloid-β (Aβ) causes more widespread neuronal dysfunction. Specific anticonvulsant therapy reverses memory impairments in various transgenic mouse strains, but it is not known whether reduction of epileptiform activity might serve as a surrogate marker of drug efficacy for memory improvement in AD mouse models.MethodsTransgenic AD mice (APP/PS1 and 3xTg-AD) were chronically implanted with dural electroencephalography electrodes, and epileptiform activity was correlated with spatial memory function and transgene-specific pathology. The antiepileptic drugs ethosuximide and brivaracetam were tested for their ability to suppress epileptiform activity and to reverse memory impairments and synapse loss in APP/PS1 mice.ResultsWe report that in two transgenic mouse models of AD (APP/PS1 and 3xTg-AD), the presence of spike-wave discharges (SWDs) correlated with impairments in spatial memory. Both ethosuximide and brivaracetam reduce mouse SWDs, but only brivaracetam reverses memory impairments in APP/PS1 mice.ConclusionsOur data confirm an intriguing therapeutic role of anticonvulsant drugs targeting synaptic vesicle protein 2A across AD mouse models. Chronic ethosuximide dosing did not reverse spatial memory impairments in APP/PS1 mice, despite reduction of SWDs. Our data indicate that SWDs are not a reliable surrogate marker of appropriate target engagement for reversal of memory dysfunction in APP/PS1 mice.

Early Activation of Experience-Independent Dendritic Spine Turnover in a Mouse Model of Alzheimer's Disease

Synaptic loss is critical in Alzheimers disease (AD), but the dynamics of synapse turnover are poorly defined. We imaged dendritic spines in transgenic APPswe/PSen1∆E9 (APP/PS1) cerebral cortex. Dendritic spine turnover is increased far from plaque in aged APP/PS1 mice, and in young APP/PS1 mice prior to plaque formation. Dysregulation occurs in the presence of soluble Aβ oligomer and requires cellular prion protein (PrPC). APP/PS1 mice lack responsiveness of spine turnover to sensory stimulation. Critically, enhanced spine turnover is coupled with the loss of persistent spines starting early and continuing with age. To evaluate mechanisms of experience-independent supranormal spine turnover, we analyzed the transcriptome of young APP/PS1 mouse brain when turnover is altered but synapse density and memory are normal, and plaque and inflammation are absent. Early PrPC-dependent expression changes occur in synaptic and lipid-metabolizing genes. Thus, pathologic synaptic dysregulation underlying AD begins at a young age prior to Aβ plaque.

Opposing effects of progranulin deficiency on amyloid and tau pathologies via microglial TYROBP network

Progranulin (PGRN) is implicated in Alzheimer’s disease (AD) as well as frontotemporal lobar degeneration. Genetic studies demonstrate an association of the common GRN rs5848 variant that results in reduced PGRN levels with increased risk for AD. However, the mechanisms by which PGRN reduction from the GRN AD risk variant or mutation exacerbates AD pathophysiology remain ill defined. Here, we show that the GRN AD risk variant has no significant effects on florbetapir positron emission tomographic amyloid imaging and cerebrospinal fluid (CSF) Aβ levels, whereas it is associated with increased CSF tau levels in human subjects of the Alzheimer’s disease neuroimaging initiative studies. Consistent with the human data, subsequent analyses using the APPswe/PS1ΔE9 (APP/PS1) mouse model of cerebral amyloidosis show that PGRN deficiency has no exacerbating effects on Aβ pathology. In contrast and unexpectedly, PGRN deficiency significantly reduces diffuse Aβ plaque growth in these APP/PS1 mice. This protective effect is due, at least in part, to enhanced microglial Aβ phagocytosis caused by PGRN deficiency-induced expression of TYROBP network genes (TNG) including an AD risk factor Trem2. PGRN-deficient APP/PS1 mice also exhibit less severe axonal dystrophy and partially improved behavior phenotypes. While PGRN deficiency reduces these amyloidosis-related phenotypes, other neuronal injury mechanisms are increased by loss of PGRN, revealing a multidimensional interaction of GRN with AD. For example, C1q complement deposition at synapses is enhanced in APP/PS1 mice lacking PGRN. Moreover, PGRN deficiency increases tau AT8 and AT180 pathologies in human P301L tau-expressing mice. These human and rodent data suggest that global PGRN reduction induces microglial TNG expression and increases AD risk by exacerbating neuronal injury and tau pathology, rather than by accelerating Aβ pathology.

Liquid and Hydrogel Phases of PrPC Linked to Conformation Shifts and Triggered by Alzheimer’s Amyloid-β Oligomers

Protein phase separation by low-complexity, intrinsically disordered domains generates membraneless organelles and links to neurodegeneration. Cellular prion protein (PrPC) contains such domains, causes spongiform degeneration, and is a receptor for Alzheimers amyloid-β oligomers (Aβo). Here, we show that PrPC separates as a liquid phase, in which α-helical Thr become unfolded. At the cell surface, PrPC Lys residues interact with Aβo to create a hydrogel containing immobile Aβo and relatively mobile PrPC. The Aβo/PrP hydrogel has a well-defined stoichiometry and dissociates with excess Aβo. NMR studies of hydrogel PrPC reveal a distinct α-helical conformation for natively unfolded amino-terminal Gly and Ala residues. Aβo/PrP hydrogel traps signal-transducing mGluR5 on the plasma membrane. Recombinant PrPC extracts endogenous Aβo from human Alzheimers soluble brain lysates into hydrogel, and a PrPC antagonist releases Aβo from endogenous brain hydrogel. Thus, coupled phase and conformational transitions of PrPC are driven by Aβ species from Alzheimers disease.