A growing number of human neurodegenerative diseases result from the expansion of a glutamine repeat in the protein that causes the disease. Spinocerebellar ataxia type 1 (SCA1) is one such disease-caused by expansion of a polyglutamine tract in the protein ataxin-1. To elucidate the genetic pathways and molecular mechanisms underlying neuronal degeneration in this group of diseases, we have created a model system for SCA1 by expressing the full-length human SCA1 gene in Drosophila. Here we show that high levels of wild-type ataxin-1 can cause degenerative phenotypes similar to those caused by the expanded protein. We conducted genetic screens to identify genes that modify SCA1-induced neurodegeneration. Several modifiers highlight the role of protein folding and protein clearance in the development of SCA1. Furthermore, new mechanisms of polyglutamine pathogenesis were revealed by the discovery of modifiers that are involved in RNA processing, transcriptional regulation and cellular detoxification. These findings may be relevant to the treatment of polyglutamine diseases and, perhaps, to other neurodegenerative diseases, such as Alzheimer's and Parkinson's disease.

Spinocerebellar ataxia type 1 (SCA1) is one of several neurological disorders caused by a CAG repeat expansion. In SCA1, this expansion produces an abnormally long polyglutamine tract in the protein ataxin-1. Mutant polyglutamine proteins accumulate in neurons, inducing neurodegeneration, but the mechanism underlying this accumulation has been unclear. We have discovered that the 14-3-3 protein, a multifunctional regulatory molecule, mediates the neurotoxicity of ataxin-1 by binding to and stabilizing ataxin-1, thereby slowing its normal degradation. The association of ataxin-1 with 14-3-3 is regulated by Akt phosphorylation, and in a Drosophila model of SCA1, both 14-3-3 and Akt modulate neurodegeneration. Our finding that phosphatidylinositol 3-kinase/Akt signaling and 14-3-3 cooperate to modulate the neurotoxicity of ataxin-1 provides insight into SCA1 pathogenesis and identifies potential targets for therapeutic intervention.

Alzheimer's disease (AD) is an incurable neurodegenerative disorder clinically characterized by progressive cognitive impairment. A prominent pathologic hallmark in the AD brain is the abnormal accumulation of the amyloid-β 1-42 peptide (Aβ), but the exact pathways mediating Aβ neurotoxicity remain enigmatic. Endoplasmic reticulum (ER) stress is induced during AD, and has been indirectly implicated as a mediator of Aβ neurotoxicity. We report here that Aβ activates the ER stress response factor X-box binding protein 1 (XBP1) in transgenic flies and in mammalian cultured neurons, yielding its active form, the transcription factor XBP1s. XBP1s shows neuroprotective activity in two different AD models, flies expressing Aβ and mammalian cultured neurons treated with Aβ oligomers. Trying to identify the mechanisms mediating XBP1s neuroprotection, we found that in PC12 cells treated with Aβ oligomers, XBP1s prevents the accumulation of free calcium (Ca(2+)) in the cytosol. This protective activity can be mediated by the downregulation of a specific isoform of the ryanodine Ca(2+) channel, RyR3. In support of this observation, a mutation in the only ryanodine receptor (RyR) in flies also suppresses Aβ neurotoxicity, indicating the conserved mechanisms between the two AD models. These results underscore the functional relevance of XBP1s in Aβ toxicity, and uncover the potential of XBP1 and RyR as targets for AD therapeutics.

BackgroundAlzheimer's disease (AD) is an age related progressive neurodegenerative disorder. One of the reasons for Alzheimer's neuropathology is the generation of large aggregates of Aß42 that are toxic in nature and induce oxidative stress, aberrant signaling and many other cellular alterations that trigger neuronal cell death. However, the exact mechanisms leading to cell death are not clearly understood.Methodology/Principal FindingsWe employed a Drosophila eye model of AD to study how Aß42 causes cell death. Misexpression of higher levels of Aß42 in the differentiating photoreceptors of fly retina rapidly induced aberrant cellular phenotypes and cell death. We found that blocking caspase-dependent cell death initially blocked cell death but did not lead to a significant rescue in the adult eye. However, blocking the levels of c-Jun NH (2)-terminal kinase (JNK) signaling pathway significantly rescued the neurodegeneration phenotype of Aß42 misexpression both in eye imaginal disc as well as the adult eye. Misexpression of Aß42 induced transcriptional upregulation of puckered (puc), a downstream target and functional read out of JNK signaling. Moreover, a three-fold increase in phospho-Jun (activated Jun) protein levels was seen in Aß42 retina as compared to the wild-type retina. When we blocked both caspases and JNK signaling simultaneously in the fly retina, the rescue of the neurodegenerative phenotype is comparable to that caused by blocking JNK signaling pathway alone.Conclusions/SignificanceOur data suggests that (i) accumulation of Aß42 plaques induces JNK signaling in neurons and (ii) induction of JNK contributes to Aß42 mediated cell death. Therefore, inappropriate JNK activation may indeed be relevant to the AD neuropathology, thus making JNK a key target for AD therapies.

Synapses are specialized communication points between neurons, and their number is a major determinant of cognitive abilities. These dynamic structures undergo developmental-and activity-dependent changes. During brain aging and certain diseases, synapses are gradually lost, causing mental decline. It is, thus, critical to identify the molecular mechanisms controlling synapse number. We show here that the levels of phosphoinositide 3 kinase (PI3K) regulate synapse number in both Drosophila larval motor neurons and adult brain projection neurons. The supernumerary synapses induced by PI3K overexpression are functional and elicit changes in behavior. Remarkably, PI3K activation induces synaptogenesis in aged adult neurons as well. We demonstrate that persistent PI3K activity is necessary for synapse maintenance. We also report that PI3K controls the expression and localization of synaptic markers in human neuroblastoma cells, suggesting that PI3K synaptogenic activity is conserved in humans. Thus, we propose that PI3K stimulation can be applied to prevent or delay synapse loss in normal aging and in neurological disorders.

The prion protein (PrP) is best known for its association with prion diseases. However, a controversial new role for PrP in Alzheimer disease (AD) has recently emerged. In vitro studies and mouse models of AD suggest that PrP may be involved in AD pathogenesis through a highly specific interaction with amyloid-␤ (A␤42) oligomers. Immobilized recombinant human PrP (huPrP) also exhibited high affinity and specificity for A␤42 oligomers. Here we report the novel finding that aggregated forms of huPrP and A␤42 are co-purified from AD brain extracts. Moreover, an anti-PrP antibody and an agent that specifically binds to insoluble PrP (iPrP) co-precipitate insoluble A␤ from human AD brain. Finally, using peptide membrane arrays of 99 13-mer peptides that span the entire sequence of mature huPrP, two distinct types of A␤ binding sites on huPrP are identified in vitro. One specifically binds to A␤42 and the other binds to both A␤42 and A␤40. Notably, A␤42-specific binding sites are localized predominantly in the octapeptide repeat region, whereas sites that bind both A␤40 and A␤42 are mainly in the extreme N-terminal or C-terminal domains of PrP. Our study suggests that iPrP is the major PrP species that interacts with insoluble A␤42 in vivo. Although this work indicated the interaction of A␤42 with huPrP in the AD brain, the pathophysiological relevance of the iPrP/A␤42 interaction remains to be established. Alzheimer disease (AD)2 is the leading cause of dementia in the elderly and the most common neurodegenerative disorder. The underlying pathology in AD seems to be associated with the accumulation of soluble and insoluble aggregated species of the amyloid-␤ (A␤) peptide in the brain (1). However, the mechanisms underlying A␤ deposition and neurotoxicity remain poorly understood. The cellular prion protein (PrP C ) is a glycoprotein highly expressed in the brain, and best known for its association with prion diseases. These are unique neurodegenerative disorders with an infectious, sporadic or genetic etiology, and which are characterized by deposition of misfolded, pathological PrP (PrP Sc ) in the brain (2). Interestingly, a recent interpretation of early and newer observations suggests that PrP C may play a role in the pathogenesis of AD (3). Epidemiological studies suggest that the Met/Val polymorphism at residue 129 in PrP modulates the number of A␤ deposits (4). Also, pathological evidence indicates that PrP deposits often accompany A␤ plaques in AD (5-7). Moreover, transgenic mice expressing mutant amyloid precursor protein (APP) and overexpressing hamster PrP C present an exacerbated A␤ plaque burden (8). The circumstantial evidence of an association between PrP and A␤ was greatly strengthened by the recent finding that PrP was the protein that most strongly supported the binding of cells to soluble A␤42 oligomers in a screen of 225,000 murine clones (9). The authors also showed that although A␤42 oligomers suppressed long-term potentiation (LTP) in CA1 hippocampal neurons in mouse brain slices, LTP inhibitio...

Prion diseases are incurable neurodegenerative disorders in which the normal cellular prion protein (PrPC) converts into a misfolded isoform (PrPSc) with unique biochemical and structural properties that correlate with disease. In humans, prion disorders, such as Creutzfeldt-Jakob disease, present typically with a sporadic origin, where unknown mechanisms lead to the spontaneous misfolding and deposition of wild type PrP. To shed light on how wild-type PrP undergoes conformational changes and which are the cellular components involved in this process, we analyzed the dynamics of wild-type PrP from hamster in transgenic flies. In young flies, PrP demonstrates properties of the benign PrPC; in older flies, PrP misfolds, acquires biochemical and structural properties of PrPSc, and induces spongiform degeneration of brain neurons. Aged flies accumulate insoluble PrP that resists high concentrations of denaturing agents and contains PrPSc-specific conformational epitopes. In contrast to PrPSc from mammals, PrP is proteinase-sensitive in flies. Thus, wild-type PrP rapidly converts in vivo into a neurotoxic, protease-sensitive isoform distinct from prototypical PrPSc. Next, we investigated the role of molecular chaperones in PrP misfolding in vivo. Remarkably, Hsp70 prevents the accumulation of PrPSc-like conformers and protects against PrP-dependent neurodegeneration. This protective activity involves the direct interaction between Hsp70 and PrP, which may occur in active membrane microdomains such as lipid rafts, where we detected Hsp70. These results highlight the ability of wild-type PrP to spontaneously convert in vivo into a protease-sensitive isoform that is neurotoxic, supporting the idea that protease-resistant PrPSc is not required for pathology. Moreover, we identify a new role for Hsp70 in the accumulation of misfolded PrP. Overall, we provide new insight into the mechanisms of spontaneous accumulation of neurotoxic PrP and uncover the potential therapeutic role of Hsp70 in treating these devastating disorders.

Spinocerebellar Ataxia type 1 (SCA1) and Huntington's disease (HD) are two polyglutamine disorders caused by expansion of a CAG repeat within the coding regions of the Ataxin-1 and Huntingtin proteins, respectively. While protein folding and turnover have been implicated in polyglutamine disorders in general, many clinical and pathological differences suggest that there are also disease-specific mechanisms. Taking advantage of a collection of genetic modifiers of expanded Ataxin-1-induced neurotoxicity, we performed a comparative analysis in Drosophila models of the two diseases. We show that while some modifier genes function similarly in SCA1 and HD Drosophila models, others have model-specific effects. Surprisingly, certain modifier genes modify SCA1 and HD models in opposite directions, i.e. they behave as suppressors in one case and enhancers in the other. Furthermore, we find that modulation of toxicity does not correlate with alterations in the formation of neuronal intranuclear inclusions. Our results point to potential common therapeutic targets in novel pathways, and to genes and pathways responsible for differences between Ataxin-1 and Huntingtin-induced neurodegeneration.