Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 20 de 62
Filter
Add more filters










Publication year range
1.
Int J Mol Sci ; 25(8)2024 Apr 09.
Article in English | MEDLINE | ID: mdl-38673751

ABSTRACT

Plant-derived multitarget compounds may represent a promising therapeutic strategy for multifactorial diseases, such as Alzheimer's disease (AD). Artemisinin and its derivatives were indicated to beneficially modulate various aspects of AD pathology in different AD animal models through the regulation of a wide range of different cellular processes, such as energy homeostasis, apoptosis, proliferation and inflammatory pathways. In this review, we aimed to provide an up-to-date overview of the experimental evidence documenting the neuroprotective activities of artemi-sinins to underscore the potential of these already-approved drugs for treating AD also in humans and propose their consideration for carefully designed clinical trials. In particular, the benefits to the main pathological hallmarks and events in the pathological cascade throughout AD development in different animal models of AD are summarized. Moreover, dose- and context-dependent effects of artemisinins are noted.


Subject(s)
Alzheimer Disease , Artemisinins , Neuroprotective Agents , Alzheimer Disease/drug therapy , Alzheimer Disease/metabolism , Artemisinins/therapeutic use , Artemisinins/pharmacology , Artemisinins/chemistry , Humans , Animals , Neuroprotective Agents/therapeutic use , Neuroprotective Agents/pharmacology , Disease Models, Animal , Apoptosis/drug effects
2.
Cells ; 12(19)2023 10 04.
Article in English | MEDLINE | ID: mdl-37830617

ABSTRACT

The amyloid precursor protein (APP) is a key molecular component of Alzheimer's disease (AD) pathogenesis. Proteolytic APP processing generates various cleavage products, including extracellular amyloid beta (Aß) and the cytoplasmic APP intracellular domain (AICD). Although the role of AICD in the activation of kinase signaling pathways is well established in the context of full-length APP, little is known about intracellular effects of the AICD fragment, particularly within discrete neuronal compartments. Deficits in fast axonal transport (FAT) and axonopathy documented in AD-affected neurons prompted us to evaluate potential axon-autonomous effects of the AICD fragment for the first time. Vesicle motility assays using the isolated squid axoplasm preparation revealed inhibition of FAT by AICD. Biochemical experiments linked this effect to aberrant activation of selected axonal kinases and heightened phosphorylation of the anterograde motor protein conventional kinesin, consistent with precedents showing phosphorylation-dependent regulation of motors proteins powering FAT. Pharmacological inhibitors of these kinases alleviated the AICD inhibitory effect on FAT. Deletion experiments indicated this effect requires a sequence encompassing the NPTY motif in AICD and interacting axonal proteins containing a phosphotyrosine-binding domain. Collectively, these results provide a proof of principle for axon-specific effects of AICD, further suggesting a potential mechanistic framework linking alterations in APP processing, FAT deficits, and axonal pathology in AD.


Subject(s)
Alzheimer Disease , Amyloid beta-Protein Precursor , Humans , Amyloid beta-Protein Precursor/metabolism , Amyloid beta-Peptides/metabolism , Axonal Transport , Alzheimer Disease/metabolism , Axons/metabolism
3.
Cell Biosci ; 13(1): 141, 2023 Aug 02.
Article in English | MEDLINE | ID: mdl-37533067

ABSTRACT

BACKGROUND: The amyloid precursor protein (APP), a key player in Alzheimer's disease (AD), is part of a larger gene family, including the APP like proteins APLP1 and APLP2. They share similar structures, form homo- and heterotypic dimers and exhibit overlapping functions. RESULTS: We investigated complex formation of the APP family members via two inducible dimerization systems, the FKBP-rapamycin based dimerization as well as cysteine induced dimerization, combined with co-immunoprecipitations and Blue Native (BN) gel analyses. Within the APP family, APLP1 shows the highest degree of dimerization and high molecular weight (HMW) complex formation. Interestingly, only about 20% of APP is dimerized in cultured cells whereas up to 50% of APP is dimerized in mouse brains, independent of age and splice forms. Furthermore, we could show that dimerized APP originates mostly from neurons and is enriched in synaptosomes. Finally, BN gel analysis of human cortex samples shows a significant decrease of APP dimers in AD patients compared to controls. CONCLUSIONS: Together, we suggest that loss of full-length APP dimers might correlate with loss of synapses in the process of AD.

4.
Acta Neuropathol Commun ; 11(1): 87, 2023 06 01.
Article in English | MEDLINE | ID: mdl-37259128

ABSTRACT

The amyloid precursor protein (APP) is a key player in Alzheimer`s disease (AD) and the precursor of the Aß peptide, which is generated by consecutive cleavages of ß- and γ-secretases. Familial Alzheimer's disease (FAD) describes a hereditary subgroup of AD that represents a low percentage of AD cases with an early onset of the disease. Different APP FAD mutations are thought to have qualitatively different effects on its proteolytic conversion. However, few studies have explored the pathogenic and putative physiological differences in more detail. Here, we compared different FAD mutations, located at the ß- (Swedish), α- (Flemish, Arctic, Iowa) or γ-secretase (Iberian) cleavage sites. We examined heterologous expression of APP WT and FAD mutants in non-neuronal cells and their impact on presynaptic differentiation in contacting axons of co-cultured neurons. To decipher the underlying molecular mechanism, we tested the subcellular localization, the endocytosis rate and the proteolytic processing in detail by immunoprecipitation-mass spectrometry. Interestingly, we found that only the Iberian mutation showed altered synaptogenic function. Furthermore, the APP Iowa mutant shows significantly decreased α-secretase processing which is in line with our results that APP carrying the Iowa mutation was significantly increased in early endosomes. However, most interestingly, immunoprecipitation-mass spectrometry analysis revealed that the amino acid substitutions of APP FAD mutants have a decisive impact on their processing reflected in altered Aß profiles. Importantly, N-terminally truncated Aß peptides starting at position 5 were detected preferentially for APP Flemish, Arctic, and Iowa mutants containing amino acid substitutions around the α-secretase cleavage site. The strongest change in the ratio of Aß40/Aß42 was observed for the Iberian mutation while APP Swedish showed a substantial increase in Aß1-17 peptides. Together, our data indicate that familial AD mutations located at the α-, ß-, and γ-secretase cleavage sites show considerable differences in the underlying pathogenic mechanisms.


Subject(s)
Alzheimer Disease , Amyloid beta-Protein Precursor , Humans , Amyloid beta-Protein Precursor/genetics , Amyloid beta-Protein Precursor/metabolism , Alzheimer Disease/genetics , Alzheimer Disease/metabolism , Amyloid Precursor Protein Secretases/genetics , Amyloid Precursor Protein Secretases/metabolism , Amyloid beta-Peptides/metabolism , Peptide Fragments/metabolism , Mutation/genetics , Presenilin-1/genetics
6.
Int J Mol Sci ; 24(5)2023 Feb 27.
Article in English | MEDLINE | ID: mdl-36902054

ABSTRACT

Alzheimer's disease (AD) is characterized by synaptic failure and neuronal loss. Recently, we demonstrated that artemisinins restored the levels of key proteins of inhibitory GABAergic synapses in the hippocampus of APP/PS1 mice, a model of cerebral amyloidosis. In the present study, we analyzed the protein levels and subcellular localization of α2 and α3 subunits of GlyRs, indicated as the most abundant receptor subtypes in the mature hippocampus, in early and late stages of AD pathogenesis, and upon treatment with two different doses of artesunate (ARS). Immunofluorescence microscopy and Western blot analysis demonstrated that the protein levels of both α2 and α3 GlyRs are considerably reduced in the CA1 and the dentate gyrus of 12-month-old APP/PS1 mice when compared to WT mice. Notably, treatment with low-dose ARS affected GlyR expression in a subunit-specific way; the protein levels of α3 GlyR subunits were rescued to about WT levels, whereas that of α2 GlyRs were not affected significantly. Moreover, double labeling with a presynaptic marker indicated that the changes in GlyR α3 expression levels primarily involve extracellular GlyRs. Correspondingly, low concentrations of artesunate (≤1 µM) also increased the extrasynaptic GlyR cluster density in hAPPswe-transfected primary hippocampal neurons, whereas the number of GlyR clusters overlapping presynaptic VIAAT immunoreactivities remained unchanged. Thus, here we provide evidence that the protein levels and subcellular localization of α2 and α3 subunits of GlyRs show regional and temporal alterations in the hippocampus of APP/PS1 mice that can be modulated by the application of artesunate.


Subject(s)
Alzheimer Disease , Antimalarials , Artesunate , Hippocampus , Receptors, Glycine , Animals , Mice , Alzheimer Disease/drug therapy , Alzheimer Disease/metabolism , Artesunate/therapeutic use , Hippocampus/metabolism , Receptors, Glycine/metabolism , Synapses/metabolism , Antimalarials/therapeutic use , Disease Models, Animal
7.
Aging Cell ; 22(3): e13778, 2023 03.
Article in English | MEDLINE | ID: mdl-36704841

ABSTRACT

N-methyl-D-aspartate receptors (NMDARs) are critical for the maturation and plasticity of glutamatergic synapses. In the hippocampus, NMDARs mainly contain GluN2A and/or GluN2B regulatory subunits. The amyloid precursor protein (APP) has emerged as a putative regulator of NMDARs, but the impact of this interaction to their function is largely unknown. By combining patch-clamp electrophysiology and molecular approaches, we unravel a dual mechanism by which APP controls GluN2B-NMDARs, depending on the life stage. We show that APP is highly abundant specifically at the postnatal postsynapse. It interacts with GluN2B-NMDARs, controlling its synaptic content and mediated currents, both in infant mice and primary neuronal cultures. Upon aging, the APP amyloidogenic-derived C-terminal fragments, rather than APP full-length, contribute to aberrant GluN2B-NMDAR currents. Accordingly, we found that the APP processing is increased upon aging, both in mice and human brain. Interfering with stability or production of the APP intracellular domain normalized the GluN2B-NMDARs currents. While the first mechanism might be essential for synaptic maturation during development, the latter could contribute to age-related synaptic impairments.


Subject(s)
Amyloid beta-Protein Precursor , Receptors, N-Methyl-D-Aspartate , Mice , Humans , Animals , Amyloid beta-Protein Precursor/genetics , Amyloid beta-Protein Precursor/metabolism , Receptors, N-Methyl-D-Aspartate/metabolism , Signal Transduction , Hippocampus/metabolism , Synapses/metabolism
8.
Cell Rep ; 41(9): 111710, 2022 11 29.
Article in English | MEDLINE | ID: mdl-36450258

ABSTRACT

The precise regulation of synaptic connectivity and function is essential to maintain neuronal circuits. Here, we show that the Drosophila pseudo-kinase Madm/NRBP1 (Mlf-1-adapter-molecule/nuclear-receptor-binding protein 1) is required presynaptically to maintain synaptic stability and to coordinate synaptic growth and function. Presynaptic Madm mediates these functions by controlling cap-dependent translation via the target of rapamycin (TOR) effector 4E-BP/Thor (eukaryotic initiation factor 4E binding protein/Thor). Strikingly, at degenerating neuromuscular synapses, postsynaptic Madm induces a compensatory, transsynaptic signal that utilizes the presynaptic homeostatic potentiation (PHP) machinery to offset synaptic release deficits and to delay synaptic degeneration. Madm is not required for canonical PHP but induces a neurodegeneration-specific form of PHP and acts via the regulation of the cap-dependent translation regulators 4E-BP/Thor and S6-kinase. Consistently, postsynaptic induction of canonical PHP or TOR activation can compensate for postsynaptic Madm to alleviate functional and structural synaptic defects. Our results provide insights into the molecular mechanisms underlying neurodegeneration-induced PHP with potential neurotherapeutic applications.


Subject(s)
Drosophila , Oligonucleotides , Animals , Homeostasis , Sirolimus/pharmacology , Software
9.
Small ; 18(47): e2202492, 2022 11.
Article in English | MEDLINE | ID: mdl-36228092

ABSTRACT

Membrane proteins can be examined in near-native lipid-bilayer environments with the advent of polymer-encapsulated nanodiscs. These nanodiscs self-assemble directly from cellular membranes, allowing in vitro probing of membrane proteins with techniques that have previously been restricted to soluble or detergent-solubilized proteins. Often, however, the high charge densities of existing polymers obstruct bioanalytical and preparative techniques. Thus, the authors aim to fabricate electroneutral-yet water-soluble-polymer nanodiscs. By attaching a sulfobetaine group to the commercial polymers DIBMA and SMA(2:1), these polyanionic polymers are converted to the electroneutral maleimide derivatives, Sulfo-DIBMA and Sulfo-SMA(2:1). Sulfo-DIBMA and Sulfo-SMA(2:1) readily extract proteins and phospholipids from artificial and cellular membranes to form nanodiscs. Crucially, the electroneutral nanodiscs avert unspecific interactions, thereby enabling new insights into protein-lipid interactions through lab-on-a-chip detection and in vitro translation of membrane proteins. Finally, the authors create a library comprising thousands of human membrane proteins and use proteome profiling by mass spectrometry to show that protein complexes are preserved in electroneutral nanodiscs.


Subject(s)
Lipid Bilayers , Nanostructures , Humans , Lipid Bilayers/chemistry , Polymers/chemistry , Maleates/chemistry , Membrane Proteins/chemistry , Nanostructures/chemistry
10.
Autophagy ; 18(9): 2068-2085, 2022 09.
Article in English | MEDLINE | ID: mdl-34964690

ABSTRACT

PSENEN/PEN2 is the smallest subunit of the γ-secretase complex, an intramembrane protease that cleaves proteins within their transmembrane domains. Mutations in components of the γ-secretase underlie familial Alzheimer disease. In addition to its proteolytic activity, supplementary, γ-secretase independent, functions in the macroautophagy/autophagy-lysosome system have been proposed. Here, we screened for PSENEN-interacting proteins and identified CLN3. Mutations in CLN3 are causative for juvenile neuronal ceroid lipofuscinosis, a rare lysosomal storage disorder considered the most common neurodegenerative disease in children. As mutations in the PSENEN and CLN3 genes cause different neurodegenerative diseases, understanding shared cellular functions of both proteins might be pertinent for understanding general cellular mechanisms underlying neurodegeneration. We hypothesized that CLN3 modulates γ-secretase activity and that PSENEN and CLN3 play associated roles in the autophagy-lysosome system. We applied CRISPR gene-editing and obtained independent isogenic HeLa knockout cell lines for PSENEN and CLN3. Following previous studies, we demonstrate that PSENEN is essential for forming a functional γ-secretase complex and is indispensable for γ-secretase activity. In contrast, CLN3 does not modulate γ-secretase activity to a significant degree. We observed in PSENEN- and CLN3-knockout cells corresponding alterations in the autophagy-lysosome system. These include reduced activity of lysosomal enzymes and lysosome number, an increased number of autophagosomes, increased lysosome-autophagosome fusion, and elevated levels of TFEB (transcription factor EB). Our study strongly suggests converging roles of PSENEN and CLN3 in the autophagy-lysosome system in a γ-secretase activity-independent manner, supporting the idea of common cytopathological processes underlying different neurodegenerative diseases.Abbreviations: Aß, amyloid-beta; AD, Alzheimer disease; APP, amyloid precursor protein; ATP5MC, ATP synthase membrane subunit c; DQ-BSA, dye-quenched bovine serum albumin; ER, endoplasmic reticulum; GFP, green fluorescent protein; ICC, immunocytochemistry; ICD, intracellular domain; JNCL, juvenile neuronal ceroid lipofuscinosis; KO, knockout; LC3, microtubule associated protein 1 light chain 3; NCL, neuronal ceroid lipofuscinoses; PSEN, presenilin; PSENEN/PEN2: presenilin enhancer, gamma-secretase subunit; TAP, tandem affinity purification; TEV, tobacco etch virus; TF, transferrin; WB, Western blot; WT, wild type.


Subject(s)
Alzheimer Disease , Neuronal Ceroid-Lipofuscinoses , Alzheimer Disease/genetics , Alzheimer Disease/metabolism , Amyloid Precursor Protein Secretases/genetics , Amyloid Precursor Protein Secretases/metabolism , Autophagy/genetics , Child , Humans , Lysosomes/metabolism , Membrane Glycoproteins/genetics , Membrane Glycoproteins/metabolism , Membrane Proteins/metabolism , Molecular Chaperones/metabolism , Neuronal Ceroid-Lipofuscinoses/genetics , Neuronal Ceroid-Lipofuscinoses/metabolism , Presenilins/genetics , Presenilins/metabolism , Transcription Factors/metabolism
11.
Biol Chem ; 403(1): 73-87, 2022 01 26.
Article in English | MEDLINE | ID: mdl-33878252

ABSTRACT

Artemisinins, a group of plant-derived sesquiterpene lactones, are efficient antimalarial agents. They also share anti-inflammatory and anti-viral activities and were considered for treatment of neurodegenerative disorders like Alzheimer's disease (AD). Additionally, artemisinins bind to gephyrin, the multifunctional scaffold of GABAergic synapses, and modulate inhibitory neurotransmission in vitro. We previously reported an increased expression of gephyrin and GABAA receptors in early pre-symptomatic stages of an AD mouse model (APP-PS1) and in parallel enhanced CDK5-dependent phosphorylation of gephyrin at S270. Here, we studied the effects of artemisinin on gephyrin in the brain of young APP-PS1 mice. We detected an additional increase of gephyrin protein level, elevated gephyrin phosphorylation at Ser270, and an increased amount of GABAAR-γ2 subunits after artemisinin-treatment. Interestingly, the CDK5 activator p35 was also upregulated. Moreover, we demonstrate decreased density of postsynaptic gephyrin and GABAAR-γ2 immunoreactivities in cultured hippocampal neurons expressing gephyrin with alanine mutations at two CDK5 phosphorylation sites. In addition, the activity-dependent modulation of synaptic protein density was abolished in neurons expressing gephyrin lacking one or both of these phosphorylation sites. Thus, our results reveal that artemisinin modulates expression as well as phosphorylation of gephyrin at sites that might have important impact on GABAergic synapses in AD.


Subject(s)
Artemisinins , Carrier Proteins , Membrane Proteins , Animals , Artemisinins/pharmacology , Carrier Proteins/metabolism , Cells, Cultured , Hippocampus/metabolism , Mice , Phosphorylation , Receptors, GABA-A/metabolism , Synapses/metabolism , gamma-Aminobutyric Acid/metabolism
12.
Biol Chem ; 403(1): 43-71, 2022 01 26.
Article in English | MEDLINE | ID: mdl-34619027

ABSTRACT

Brain-derived neurotrophic factor (BDNF) is an important modulator for a variety of functions in the central nervous system (CNS). A wealth of evidence, such as reduced mRNA and protein level in the brain, cerebrospinal fluid (CSF), and blood samples of Alzheimer's disease (AD) patients implicates a crucial role of BDNF in the progression of this disease. Especially, processing and subcellular localization of BDNF and its receptors TrkB and p75 are critical determinants for survival and death in neuronal cells. Similarly, the amyloid precursor protein (APP), a key player in Alzheimer's disease, and its cleavage fragments sAPPα and Aß are known for their respective roles in neuroprotection and neuronal death. Common features of APP- and BDNF-signaling indicate a causal relationship in their mode of action. However, the interconnections of APP- and BDNF-signaling are not well understood. Therefore, we here discuss dimerization properties, localization, processing by α- and γ-secretase, relevance of the common interaction partners TrkB, p75, sorLA, and sortilin as well as shared signaling pathways of BDNF and sAPPα.


Subject(s)
Alzheimer Disease , Brain-Derived Neurotrophic Factor , ADAM10 Protein , Adaptor Proteins, Vesicular Transport , Amyloid Precursor Protein Secretases , Amyloid beta-Peptides , Amyloid beta-Protein Precursor , Humans , LDL-Receptor Related Proteins , Membrane Glycoproteins , Membrane Proteins , Membrane Transport Proteins , Nerve Tissue Proteins , Receptor, trkB , Receptors, Nerve Growth Factor
13.
Biol Chem ; 403(1): 3-26, 2022 01 26.
Article in English | MEDLINE | ID: mdl-34449171

ABSTRACT

Mycotoxins are fungal metabolites that can cause various diseases in humans and animals. The adverse health effects of mycotoxins such as liver failure, immune deficiency, and cancer are well-described. However, growing evidence suggests an additional link between these fungal metabolites and neurodegenerative diseases. Despite the wealth of these initial reports, reliable conclusions are still constrained by limited access to human patients and availability of suitable cell or animal model systems. This review summarizes knowledge on mycotoxins associated with neurodegenerative diseases and the assumed underlying pathophysiological mechanisms. The limitations of the common in vivo and in vitro experiments to identify the role of mycotoxins in neurotoxicity and thereby in neurodegenerative diseases are elucidated and possible future perspectives to further evolve this research field are presented.


Subject(s)
Mycotoxins , Neurodegenerative Diseases , Animals , Fungi , Humans , Neurodegenerative Diseases/drug therapy
15.
Cells ; 10(7)2021 06 25.
Article in English | MEDLINE | ID: mdl-34202290

ABSTRACT

The scaffolding protein family Fe65, composed of Fe65, Fe65L1, and Fe65L2, was identified as an interaction partner of the amyloid precursor protein (APP), which plays a key function in Alzheimer's disease. All three Fe65 family members possess three highly conserved interaction domains, forming complexes with diverse binding partners that can be assigned to different cellular functions, such as transactivation of genes in the nucleus, modulation of calcium homeostasis and lipid metabolism, and regulation of the actin cytoskeleton. In this article, we rule out putative new intracellular signaling mechanisms of the APP-interacting protein Fe65 in the regulation of actin cytoskeleton dynamics in the context of various neuronal functions, such as cell migration, neurite outgrowth, and synaptic plasticity.


Subject(s)
Actins/metabolism , Nerve Tissue Proteins/metabolism , Actin Cytoskeleton/metabolism , Amyloid beta-Protein Precursor/metabolism , Animals , Humans , Models, Biological , Nerve Tissue Proteins/genetics , Protein Binding
16.
Mol Cell Neurosci ; 113: 103624, 2021 06.
Article in English | MEDLINE | ID: mdl-33933588

ABSTRACT

Alzheimer's disease (AD) is the most frequent form of dementia, characterized histopathologically by the formation of amyloid plaques and neurofibrillary tangles in the brain. Amyloid ß-peptide (Aß) is a major component of amyloid plaques and is released together with carboxy-terminal fragments (CTFs) from the amyloid precursor protein (APP) through proteolytic cleavage, thought to contribute to synapse dysfunction and loss along the progression of AD. Artemisinins, primarily antimalarial drugs, reduce neuroinflammation and improve cognitive capabilities in mouse models of AD. Furthermore, artemisinins were demonstrated to target gephyrin, the main scaffold protein of inhibitory synapses and modulate GABAergic neurotransmission in vitro. Previously, we reported a robust decrease of inhibitory synapse proteins in the hippocampus of 12-month-old double transgenic APP-PS1 mice which overexpress in addition to the Swedish mutated form of the human APP a mutated presenilin 1 (PS1) gene and are characterized by a high plaque load at this age. Here, we provide in vivo evidence that treating these mice with artemisinin or its semisynthetic derivative artesunate in two different doses (10 mg/kg and 100 mg/kg), these compounds affect differently inhibitory synapse components, amyloid plaque load and APP-processing. Immunofluorescence microscopy demonstrated the rescue of gephyrin and γ2-GABAA-receptor protein levels in the brain of treated mice with both, artemisinin and artesunate, most efficiently with a low dose of artesunate. Remarkably, artemisinin reduced only in low dose the amyloid plaque load correlating with lower levels of mutated human APP (hAPPswe) whereas artesunate treatment in both doses resulted in significantly lower plaque numbers. Correspondingly, the level of APP-cleavage products, specifically the amount of CTFs in hippocampus homogenates was reduced significantly only by artesunate, in line with the findings in hAPPswe expressing cultured hippocampal neurons evidencing a concentration-dependent inhibition of CTF-release by artesunate already in the nanomolar range. Thus, our data support artemisinins as neuroprotective multi-target drugs, exhibiting a potent anti-amyloidogenic activity and reinforcing key proteins of inhibitory synapses.


Subject(s)
Alzheimer Disease/drug therapy , Artesunate/therapeutic use , Neuroprotective Agents/therapeutic use , Synapses/metabolism , Alzheimer Disease/metabolism , Amyloid beta-Protein Precursor/metabolism , Animals , Artesunate/pharmacology , Cells, Cultured , Hippocampus/cytology , Hippocampus/drug effects , Hippocampus/metabolism , Male , Membrane Proteins/metabolism , Mice , Mice, Inbred C57BL , Neurons/drug effects , Neurons/metabolism , Neuroprotective Agents/pharmacology , Receptors, GABA-A/metabolism , Synapses/drug effects
17.
J Alzheimers Dis ; 74(4): 1167-1187, 2020.
Article in English | MEDLINE | ID: mdl-32144981

ABSTRACT

Early changes in inhibitory synapse connectivities are thought to contribute to the excitation/inhibition imbalance preceding neurodegeneration in Alzheimer's disease (AD). Recently, we reported a robust increase in the level of different key-proteins of inhibitory synapses in hippocampal subregions of pre-symptomatic APPswe-PS1 mice, a model of cerebral amyloidosis. Besides increased inhibitory synaptic clusters on parvalbumin-positive projections in CA1 and CA3, we observed impaired communication between these two hippocampal areas of young APP-PS1 mice. Interestingly, the phosphorylation of gephyrin, a major organizer of inhibitory synapses, was also increased. Here, we demonstrate that the protein levels of CDK5, a kinase involved in the phosphorylation of gephyrin, and its regulatory protein p35 are also significantly increased in hippocampal subregions of young APP-PS1 mice. Consistently, the expression of hAPP-swe in cultured hippocampal neurons resulted in higher p35-protein levels, indicating a possible molecular link between increased Aß-production and the elevated p35/CDK5 levels seen in vivo. Further, a shRNA mediated downregulation of p35-expression in hippocampal neurons correlated with a decrease in gephyrin phosphorylation and in a reduced density of synaptic γ2-GABAA-receptor clusters. These findings, together with the detection of gephyrin colocalization with CDK5 and p35 by immunostaining and proximity ligation experiments in vivo and in vitro, are supporting our hypothesis that Aß has a profound impact on inhibitory network properties, likely mediated at least in part by p35/CDK5 signaling. This further underscores the impact of altered inhibitory synaptic transmission in AD.


Subject(s)
Amyloid Neuropathies/metabolism , Amyloid beta-Peptides/metabolism , Cyclin-Dependent Kinase 5/metabolism , Phosphotransferases/metabolism , Signal Transduction , Synapses/physiology , Amyloid Neuropathies/physiopathology , Animals , Brain/metabolism , Brain/physiopathology , Cells, Cultured , Disease Models, Animal , Gene Knockdown Techniques , Hippocampus/metabolism , Hippocampus/physiopathology , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Rats , Synapses/metabolism
18.
Mol Biol Rep ; 47(4): 3019-3024, 2020 Apr.
Article in English | MEDLINE | ID: mdl-32152789

ABSTRACT

Glyceraldehyde 3-phosphate dehydrogenase's (GAPDH) proapoptotic response to cellular oxidative stress has suspected implication for Alzheimer's disease (AD). Interestingly, the overexpression of the amyloid precursor protein (APP) can initiate oxidative stress responses within mammalian cell lines. Here, APP695 and APP770 overexpression significantly increased the level of GAPDH, while no effect was observed when the APP homologues APLP1 or APLP2 were used. Heterologous expression of APP695 was shown to increase the level of GAPDH within the cytoplasm by over 100% and within the mitochondria by approximately 50%. Moreover, a shift in organelle distribution from cytoplasm > nucleus > mitochondria in control cell lines to cytoplasm > mitochondria > nucleus in the APP695 overexpressing cell line was also observed. Further, the overexpression of APP695 increased GAPDH aggregation temperature by 3.09 ± 0.46 °C, indicative of greater thermal stability. These results demonstrate a clear correlation between APP overexpression and GAPDH levels, organelle distribution and thermal stability.


Subject(s)
Amyloid beta-Protein Precursor/metabolism , Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating)/metabolism , Oxidative Stress/physiology , Alzheimer Disease/metabolism , Amyloid beta-Protein Precursor/physiology , Cytoplasm/metabolism , Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating)/physiology , Glyceraldehyde-3-Phosphate Dehydrogenases/metabolism , Glyceraldehyde-3-Phosphate Dehydrogenases/physiology , HEK293 Cells , Humans , Mitochondria/metabolism , Oxidation-Reduction
19.
Cell Mol Life Sci ; 77(24): 5223-5242, 2020 Dec.
Article in English | MEDLINE | ID: mdl-32065241

ABSTRACT

Endocytosis of the amyloid precursor protein (APP) is critical for generation of ß-amyloid, aggregating in Alzheimer's disease. APP endocytosis depending on the intracellular NPTY motif is well investigated, whereas involvement of the YTSI (also termed BaSS) motif remains controversial. Here, we show that APP lacking the YTSI motif (ΔYTSI) displays reduced localization to early endosomes and decreased internalization rates, similar to APP ΔNPTY. Additionally, we show that the YTSI-binding protein, PAT1a interacts with the Rab5 activator RME-6, as shown by several independent assays. Interestingly, knockdown of RME-6 decreased APP endocytosis, whereas overexpression increased the same. Similarly, APP ΔNPTY endocytosis was affected by PAT1a and RME-6 overexpression, whereas APP ΔYTSI internalization remained unchanged. Moreover, we could show that RME-6 mediated increase of APP endocytosis can be diminished upon knocking down PAT1a. Together, our data identify RME-6 as a novel player in APP endocytosis, involving the YTSI-binding protein PAT1a.


Subject(s)
Alzheimer Disease/genetics , Amino Acid Motifs/genetics , Amyloid beta-Protein Precursor/genetics , rab5 GTP-Binding Proteins/genetics , Alzheimer Disease/pathology , Amyloid beta-Peptides/genetics , Animals , Carrier Proteins/genetics , Endocytosis/genetics , Endosomes/genetics , Humans , Mice , Protein Transport/genetics , Transport Vesicles/genetics
20.
J Neurochem ; 151(5): 626-641, 2019 12.
Article in English | MEDLINE | ID: mdl-31063592

ABSTRACT

The amyloid precursor protein (APP) and its homologs amyloid precursor-like protein 1 (APLP1) and APLP2 have central physiological functions in transcellular adhesion that depend on copper and zinc mediated trans-directed dimerization of the extracellular domains E1 and E2. Copper binds to three distinct sites in APP, one in the copper binding (CuBD) and growth factor-like (GFLD) domains each within E1, and one in the E2 domain. For APLP1 and APLP2, metal binding has so far only been shown for the E2 domain. Zinc binding has been reported for all APP family members to a unique site in the E2 domain and an additional site essential for APLP1 E2 domain trans-dimerization. Using isothermal titration calorimetry, co-immunoprecipitation, and in vitro bead aggregation assays, we show that copper promotes cis- as well as trans-directed dimerization of APLP1 and APLP2, similar as reported previously for APP. Furthermore, we report a APP-specific zinc binding site with nanomolar affinity located in the E1 domain, whereas no binding of zinc to the individual subdomains GFLD or CuBD was detected. Zinc binding did not affect the cis- but trans-dimerization of APP and APLP1. Furthermore, zinc binding inhibited copper-induced trans-directed dimerization of APP. Together, we identified a high-affinity APP-specific zinc binding site in the E1 domain and revealed contrasting cis- and trans-directed dimerization properties of APP, APLP1, and APLP2 in dependence on zinc and copper ions. Consequently, changes in metal ion homeostasis, as reported in the context of synaptic activity and neurodegenerative diseases, appear as key modulators of homo- and heterotypic trans-cellular APP/APLPs complexes.


Subject(s)
Amyloid beta-Protein Precursor/chemistry , Copper/chemistry , Protein Multimerization/physiology , Zinc/chemistry , Animals , Humans , Protein Domains
SELECTION OF CITATIONS
SEARCH DETAIL
...