Yun-wu Zhang

Differential Regulation of Amyloid-β Endocytic Trafficking and Lysosomal Degradation by Apolipoprotein E Isoforms

Background: Apolipoprotein E (apoE) regulates amyloid-β (Aβ) clearance in an isoform-dependent manner. Results: Internalized Aβ traffics to lysosomal and recycling pathways. ApoE3 more efficiently promotes Aβ lysosomal trafficking and degradation than apoE4. Conclusion: ApoE isoforms differentially affect Aβ lysosomal trafficking and degradation. Significance: Differential effects of apoE isoforms on Aβ cellular degradation may explain why apoE4 is a risk factor for Alzheimer disease. Aggregation of amyloid-β (Aβ) peptides leads to synaptic disruption and neurodegeneration in Alzheimer disease (AD). A major Aβ clearance pathway in the brain is cellular uptake and degradation. However, how Aβ traffics through the endocytic pathway and how AD risk factors regulate this event is unclear. Here we show that the majority of endocytosed Aβ in neurons traffics through early and late endosomes to the lysosomes for degradation. Overexpression of Rab5 or Rab7, small GTPases that function in vesicle fusion for early and late endosomes, respectively, significantly accelerates Aβ endocytic trafficking to the lysosomes. We also found that a portion of endocytosed Aβ traffics through Rab11-positive recycling vesicles. A blockage of this Aβ recycling pathway with a constitutively active Rab11 mutant significantly accelerates cellular Aβ accumulation. Inhibition of lysosomal enzymes results in Aβ accumulation and aggregation. Importantly, apolipoprotein E (apoE) accelerates neuronal Aβ uptake, lysosomal trafficking, and degradation in an isoform-dependent manner with apoE3 more efficiently facilitating Aβ trafficking and degradation than apoE4, a risk factor for AD. Taken together, our results demonstrate that Aβ endocytic trafficking to lysosomes for degradation is a major Aβ clearance pathway that is differentially regulated by apoE isoforms. A disturbance of this pathway can lead to accumulation and aggregation of cellular Aβ capable of causing neurotoxicity and seeding amyloid.

Estrogen, beta-amyloid metabolism/trafficking, and Alzheimer's disease.

Abstract:u2002 Estrogen plays key regulatory roles in a variety of biological actions besides its classic function as a sex hormone. Recently, estrogen has been linked to neurodegenerative diseases including Alzheimers disease (AD) and Parkinsons disease (PD). Several lines of evidence support the notion that brain estrogen exerts neuroprotective effects against various types of neurotoxicity in different cellular and animal models. Despite some controversies, estrogen replacement therapy (ERT) at an early stage, especially when given prior to menopause, has been shown to reduce the risk of AD in postmenopausal women. In addition, multiple lines of evidence have proven the neuroprotective effects of estrogen, such as enhancing neurotrophin signaling and synaptic activities pertinent to memory functions and protecting neurons against oxidative injuries and β‐amyloid toxicity; the latter is widely accepted as the prime culprit known to trigger the pathogenesis of AD. Here we will summarize our findings that estrogen decreased generation and secretion of β‐amyloid peptides in cultured cells and primary neurons and that administration of estrogen in estrogen‐deprived mice reversed the elevated levels of brain Aβ. We will also discuss the molecular and cellular mechanisms underlying estrogens effects on Aβ metabolism, which is highlighted by our demonstration that estrogen increases intracellular trafficking of β‐amyloid precursor protein (βAPP) and hence reduces maximal Aβ generation within the trans‐Golgi network (TGN), a subcellular compartment in which APP is known to be cleaved by the secretase enzymes to generate Aβ.

Appoptosin-Mediated Caspase Cleavage of Tau Contributes to Progressive Supranuclear Palsy Pathogenesis

Progressive supranuclear palsy (PSP) is a movement disorder characterized by tau neuropathology where the underlying mechanism is unknown. An SNP (rs1768208 C/T) has been identified as a strong risk factor for PSP. Here, we identified a much higher T-allele occurrence and increased levels of the pro-apoptotic protein appoptosin in PSP patients. Elevations in appoptosin correlate with activated caspase-3 and caspase-cleaved tau levels. Appoptosin overexpression increased caspase-mediated tau cleavage, tau aggregation, and synaptic dysfunction, whereas appoptosin deficiency reduced tau cleavage and aggregation. Appoptosin transduction impaired multiple motor functions and exacerbated neuropathology in tau-transgenic mice in a manner dependent on caspase-3 and tau. Increased appoptosin and caspase-3-cleaved tau were also observed in brain samples of patients with Alzheimers disease and frontotemporal dementia with tau inclusions. Our findings reveal a novel role for appoptosin in neurological disorders with tau neuropathology, linking caspase-3-mediated tau cleavage to synaptic dysfunction and behavioral/motor defects.

Transcriptional regulation of PEN-2, a key component of the gamma-secretase complex, by CREB

ABSTRACT Gamma-secretase, which is responsible for the intramembranous cleavage of Alzheimers β-amyloid precursor protein (APP), the signaling receptor Notch, and many other substrates, is a multiprotein complex consisting of at least four components: presenilin (PS), nicastrin, APH-1, and PEN-2. Despite the fact that PEN-2 is known to mediate endoproteolytic cleavage of full-length PS and APH-1 and nicastrin are required for maintaining the stability of the complex, the detailed physiological function of each component remain elusive. Unlike that of PS, the transcriptional regulation of PEN-2, APH-1, and nicastrin has not been investigated. Here, we characterized the upstream regions of the human PEN-2 gene and identified a 238-bp fragment located 353 bp upstream of the translational start codon as the key region necessary for the promoter activity. Further analysis revealed a CREB binding site located in the 238-bp region that is essential for the transcriptional activity of the PEN-2 promoter. Mutation of the CREB site abolished the transcriptional activity of the PEN-2 promoter. Electrophoretic mobility shift assays and chromatin immunoprecipitation analysis showed the binding of CREB to the PEN-2 promoter region both in vitro and in vivo. Activation of the CREB transcriptional factor by forskolin dramatically promoted the expression of PEN-2 mRNA and protein, whereas the other components of the γ-secretase complex remained unaffected. Forskolin treatment slightly increases the secretion of soluble APPα and Aβ without affecting Notch cleavage. These results demonstrate that expression of PEN-2 is regulated by CREB and suggest that the specific control of PEN-2 expression may imply additional physiological functions uniquely assigned to PEN-2.

Calnuc binds to Alzheimer's beta-amyloid precursor protein and affects its biogenesis

Calnuc, a Golgi calcium binding protein, plays a key role in the constitution of calcium storage. Abnormal calcium homeostasis has been linked to Alzheimer’s disease (AD). Excessive production and/or accumulation of β‐amyloid (Aβ) peptides that are proteolytically derived from the β‐amyloid precursor protein (APP) have been linked to the pathogenesis of AD. APP has also been indicated to play multiple physiological functions. In this study, we demonstrate that calnuc interacts with APP through direct binding to the carboxyl‐terminal region of APP, possibly in a calcium‐sensitive manner. Immunofluorescence study revealed that the two proteins co‐localize in the Golgi in both cultured cells and mouse brains. Over‐expression of calnuc in neuroblastoma cells significantly reduces the level of endogenous APP. Conversely, down‐regulation of calnuc by siRNA increases cellular levels of APP. Additionally, we show that over‐expression of calnuc down‐regulates the APP mRNA level and inhibits APP biosynthesis, which in turn results in a parallel reduction of APP proteolytic metabolites, sAPP, CTFs and Aβ. Furthermore, we found that the level of calnuc was significantly decreased in the brain of AD patients as compared with that of age‐matched non‐AD controls. Our results suggest a novel function of calnuc in modulating the levels of APP and its proteolytic metabolites, which may further affect the patho/physiological functions of APP including AD pathogenesis.

SNX27 and SORLA Interact to Reduce Amyloidogenic Subcellular Distribution and Processing of Amyloid Precursor Protein

Proteolytic generation of amyloidogenic amyloid β (Aβ) fragments from the amyloid precursor protein (APP) significantly contributes to Alzheimers disease (AD). Although amyloidogenic APP proteolysis can be affected by trafficking through genetically associated AD components such as SORLA, how SORLA functionally interacts with other trafficking components is yet unclear. Here, we report that SNX27, an endosomal trafficking/recycling factor and a negative regulator of the γ-secretase complex, binds to the SORLA cytosolic tail to form a ternary complex with APP. SNX27 enhances cell surface SORLA and APP levels in human cell lines and mouse primary neurons, and depletion of SNX27 or SORLA reduces APP endosome-to-cell surface recycling kinetics. SNX27 overexpression enhances the generation of cell surface APP cleavage products such as soluble alpha-APP C-terminal fragment (CTFα) in a SORLA-dependent manner. SORLA-mediated Aβ reduction is attenuated by downregulation of SNX27. This indicates that an SNX27/SORLA complex functionally interacts to limit APP distribution to amyloidogenic compartments, forming a non-amyloidogenic shunt to promote APP recycling to the cell surface. SIGNIFICANCE STATEMENT Many genes have been identified as risk factors for Alzheimers disease (AD), and a large proportion of these genes function to limit production or toxicity of the AD-associated amyloid β (Aβ) peptide. Whether and how these genes precisely operate to limit AD onset remains an important question. We identify binding and trafficking interactions between two of these factors, SORLA and SNX27, and demonstrate that SNX27 can direct trafficking of SORLA and the Aβ precursor APP to the cell surface to limit the production of Aβ. Diversion APP to the cell surface through modulation of this molecular complex may represent a complimentary strategy for future development in AD treatment.

Study of room-temperature phosphorescence of 1-bromonaphthalene in sodium dodecylbenzene sulfonate and beta-cyclodextrin solution

Abstract Intense room-temperature phosphorescence (RTP) of 1-bromo-naphthalene (1-BrN) was studied in aerated aqueous solutions containing sodium dodecylbenzene sulfonate (SDBS) and β-cyclodextrin (β-CD). It has been found that considerably enhanced RTP arises from the formation of the 1:1:1/SDBS:1-BrN:β-CD ternary inclusion complex in premicellar solutions. The spectral and structural analyses of molecules indicate that the phenyl ring of SDBS is included in the apolar cavity of β-CD and the polar head group and a part of hydrocarbon chain located outside the cavity. Surface tension of the solutions demonstrates that the presence of micelles results in serious phosphorescence quenching.

RPS23RG1 reduces Aβ oligomer-induced synaptic and cognitive deficits

Alzheimer’s disease (AD) is the most common form of dementia in the elderly. It is generally believed that β-amyloidogenesis, tau-hyperphosphorylation, and synaptic loss underlie cognitive decline in AD. Rps23rg1, a functional retroposed mouse gene, has been shown to reduce Alzheimer’s β-amyloid (Aβ) production and tau phosphorylation. In this study, we have identified its human homolog, and demonstrated that RPS23RG1 regulates synaptic plasticity, thus counteracting Aβ oligomer (oAβ)-induced cognitive deficits in mice. The level of RPS23RG1 mRNA is significantly lower in the brains of AD compared to non-AD patients, suggesting its potential role in the pathogenesis of the disease. Similar to its mouse counterpart, human RPS23RG1 interacts with adenylate cyclase, activating PKA/CREB, and inhibiting GSK-3. Furthermore, we show that human RPS23RG1 promotes synaptic plasticity and offsets oAβ-induced synaptic loss in a PKA-dependent manner in cultured primary neurons. Overexpression of Rps23rg1 in transgenic mice consistently prevented oAβ-induced PKA inactivation, synaptic deficits, suppression of long-term potentiation, and cognitive impairment as compared to wild type littermates. Our study demonstrates that RPS23RG1 may reduce the occurrence of key elements of AD pathology and enhance synaptic functions to counteract oAβ-induced synaptic and cognitive deficits in AD.

Substrate check of gamma-secretase.

γ-secretase cleaves multiple substrates with essential roles from development to neurodegeneration, and its aberrant processing underlies human disorders including Alzheimers disease (AD). Tian et al. now identify a domain in the β-amyloid precursor protein (APP) that inhibits γ-secretase activity and show that certain familial AD–linked APP mutations within this domain impair this inhibition, resulting in increased β-amyloid generation. The study thus reveals a novel mechanism whereby γ-secretases activity is influenced by its own substrate.

Neuron-Specific Menin Deletion Leads to Synaptic Dysfunction and Cognitive Impairment by Modulating p35 Expression

Menin (MEN1) is a critical modulator of tissue development and maintenance. As such, MEN1 mutations are associated with multiple endocrine neoplasia type 1 (MEN1) syndrome. Although menin is abundantly expressed in the nervous system, little is known with regard to its function in the adult brain. Here, we demonstrate that neuron-specific deletion of Men1 (CcKO) affects dendritic branching and spine formation, resulting in defects in synaptic function, learning, and memory. Furthermore, we find that menin binds to the p35 promoter region to facilitate p35 transcription. As a primary Cdk5 activator, p35 is expressed mainly in neurons and is critical for brain development and synaptic plasticity. Restoration of p35 expression in the hippocampus and cortex of Men1 CcKO mice rescues synaptic and cognitive deficits associated with Men1 deletion. These results reveal a critical role for menin in synaptic and cognitive function by modulating the p35-Cdk5 pathway.