Drosophila melanogaster as a model organism for

Alzheimer’s disease
Prüßing et al.

Prüßing et al. Molecular Neurodegeneration 2013, 8:35

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Prüßing et al. Molecular Neurodegeneration 2013, 8:35
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REVIEW Open Access

Drosophila melanogaster as a model organism for

Alzheimer’s disease
Katja Prüßing1, Aaron Voigt1*† and Jörg B Schulz1,2,3†

Abstract
Drosophila melanogaster provides an important resource for in vivo modifier screens of neurodegenerative diseases.
To study the underlying pathogenesis of Alzheimer’s disease, fly models that address Tau or amyloid toxicity have
been developed. Overexpression of human wild-type or mutant Tau causes age-dependent neurodegeneration,
axonal transport defects and early death. Large-scale screens utilizing a neurodegenerative phenotype induced by
eye-specific overexpression of human Tau have identified several kinases and phosphatases, apoptotic regulators
and cytoskeleton proteins as determinants of Tau toxicity in vivo. The APP ortholog of Drosophila (dAPPl) shares the
characteristic domains with vertebrate APP family members, but does not contain the human Aβ42 domain. To
circumvent this drawback, researches have developed strategies by either direct secretion of human Aβ42 or triple
transgenic flies expressing human APP, β-secretase and Drosophila γ-secretase presenilin (dPsn). Here, we provide a
brief overview of how fly models of AD have contributed to our knowledge of the pathomechanisms of disease.
Keywords: Drosophila melanogaster, Amyloid-β, Tau, Alzheimer’s disease

Background The neurofibrillary tangles are composed of hyperpho-

Alzheimer’s disease (AD) is the most common irreversible sphorylated Tau proteins and are found intracellularly in
cause of dementia. It is characterized by cognitive impair- affected neurons. In non-disease situation, Tau is bound to
ment and progressive neurodegeneration and affects more microtubuli (MT) and thereby leads to the stabilization of
than 24 million people worldwide [1]. With AD diagnoses MT. The affinity of Tau to MT is regulated by phosphoryl-
being on the rise, burdening existing healthcare support ation of Tau’s MT binding sites. A high degree of phosphor-
mechanisms, the disease is set to wreak havoc on the ylation results in detachment from MT and subsequent Tau
healthcare industry. Definite diagnosis of AD requires aggregation, finally causing the formation of neurofibrillary
the correct identification of classical neuropathological tangles [4].
hallmarks, which are extracellular amyloid plaques and The dominating, but not exclusive explanation for the
intracellular neurofibrillary tangles. molecular basis of AD pathology is the amyloid cascade
Plaques are primarily composed of Amyloid-β peptides hypothesis. It states that the deposition of Aβ in the brain
(Aβ) generated by differential proteolytic cleavage of the is the central event initiating disease progression [5]. Aβ
transmembrane receptor Amyloid Precursor Protein (APP). deposits activate downstream neurotoxic mechanisms
The endoproteolysis is performed by the β-site APP- including deregulation of Tau-MT-binding properties.
cleaving enzyme (BACE) and γ-secretases, consisting The amyloid cascade hypothesis is supported by the fact
of Presenilin 1/2, Nicastrin, APH-1 and PEN-2 [2]. Among that mutations implicated in familial AD are known to
other peptides and proteins, the two cleavage products increase ratios of Aβ42/Aβ40 and aggregation [6-8].
Aβ40 and Aβ42 are found in plaques. However, Aβ42 is Although Tau mutations lead to neurodegeneration [9],
the predominant form and is considered to be the main none of the disease-linked Tau mutations is linked to
amyloidogenic peptide as it forms fibrils more easily [3]. familial AD. Mutations in Tau rather cause fronto-temporal
dementia or progressive nuclear palsy in which Aβ42
* Correspondence: [email protected] deposits are absent [10].
†
Equal contributors
1
Department of Neurology, University Medical Center, RWTH Aachen, Several lines of evidence support the idea that Tau acts
Pauwelsstrasse 30, D-52074 Aachen, Germany downstream of Aβ42 toxicity. Clearance of Aβ reduced
Full list of author information is available at the end of the article

© 2013 Prüßing et al.; licensee BioMed Central Ltd. This is an open access article distributed under the terms of the Creative
Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and
reproduction in any medium, provided the original work is properly cited.
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early hyperphosphorylated Tau aggregation in double of well-established molecular genetics tools are available
transgenic mice, whereas increasing Tau burden did not [24]. Another advantage regarding its usefulness in
affect Aβ42 accumulation [11]. Furthermore, it is known biomedical research, especially in the field of neurode-
that reduction of Tau protein levels leads to an amelioration generative diseases, is its short lifespan. Depending on
of Aβ-induced learning and memory impairment [12]. diet and stress it ranges up to an average maximum of
Mechanisms linking extracellular Aβ42 to intracellular 120 days. All this makes Drosophila an ideal organism to
Tau are a subject of intensive research. One possible molecu- study neurodegenerative diseases like AD [25]. Previous
lar mechanism is associated with a dendritic function of studies have clearly shown that the expression of disease-
Tau [13]. Dendritic Tau targets Fyn kinase to postsynaptic related gene products (Tau protein and Aβ42 peptide,
density, where Fyn facilitates stabilization of a complex respectively) causes phenotypes in flies. Reminiscent of the
triggering downstream excitotoxic signaling [13]. situation observed in AD patients, flies show a robust de-
In modern research several model systems have been cline of neurons upon Aβ42 and/or Tau overexpression.
developed trying to reveal molecular mechanisms linking Depending on the neuronal subset the expression of the
pathological hallmarks like aggregating Tau and Aβ pep- AD-linked peptides/proteins is targeted to, the neuronal
tides to neurodegeneration finally resulting in progressive decline has different phenotypic outcomes like early death,
memory loss as observed in AD. However, key features of reduced locomotion in larvae and adults, decreased flight
the disease etiology still remain elusive and no efficient ability, blindness, rough eye texture, etc. All these parame-
therapy has been found so far. ters can be analyzed and quantified, thus making the fly
This review summarizes the utilization of Drosophila a reasonable organism to study specific aspects of AD
melanogaster to mimic AD pathology inflicted by excess pathology. In addition, more sophisticated behavioral or
Tau protein and Aβ42 peptide production. cognitive assays can be performed in flies. Applying such
assays on fly models of AD, a decline in cognition, a
Drosophila as a model organism for AD hallmark of AD was observed. Overall, the fly is a
Animal model systems are used to study specific functional powerful model to study the molecular basis of neuronal
aspects of human diseases in general and neurodegenera- decline in the context of AD [26,27]. Tests on alterations
tive diseases in particular. AD models range from yeast in behavior and/or cognition are possible in flies. However,
[14] and Caenorhabditis elegans [15] to mammals and their analysis is often time-consuming and the conclusions
human cell culture systems [16-18]. However, no model that can be drawn with regard to humans are fairly limited.
system combines easy use and essential criteria of AD, An overview of advantages and disadvantages using Dros-
like cognitive and behavioral dysfunction caused by cell ophila as a model organism to study neurodegenerative
type-specific neurodegeneration, cellular pathophysiology diseases like AD is provided in Table 1.
including aggregate formation, clear pattern of inheritance
and genetic homogeneity. Although vertebrate model Drosophila models for Aβ toxicity
organisms reflect pathologic hallmarks of human diseases Comparative analysis of whole genomes revealed striking
very well, these model organisms have the disadvantage similarities between structural composition of human and
of care, time and cost-intensive handling. Using comparable Drosophila genes [28]. Nearly 70% of human disease-
short-lived model organisms allows fast data acquisition causing genes have orthologs in the fly [29]. Given this,
facilitating large-scale experiments, although these organ- it is not surprising that orthologs associated to known
isms might lack some pathophysiological characteristics AD genes not only exist in Drosophila, but also exhibit
of AD (a summary of invertebrate AD models is pro- functional conservation.
vided in [19]). Drosophila harbors an APP ortholog [30] and all
Drosophila has more than a hundred-year history in components of the γ-secretase complex [31]. Although
genetic research [20]. It is used as prime model organism a β-secretase-like enzyme was identified in flies [32], it
for experimental studies of multi-cellular eukaryotic biology displays very low β-secretase activity [33]. The Drosophila
and it combines genetic, anatomic, behavioral, methodical APP ortholog dAPPl shares the characteristic domains with
and even economic advantages. It is one of the first organ- vertebrate APP family members [30]. However, the region
isms with a fully sequenced genome [21]. Approximately corresponding to the Aβ peptides lacks significant hom-
13,600 protein-coding genes are located in only four ology [30]. As a consequence, there is no endogenous
chromosomes. The fly anatomy is well studied, its brain Aβ production in the fly. Nevertheless, overexpression
and nervous system are quite complex [22]. Its anatomical of the β-secretase-like protein resulted in cleavage of dAPPl
features like the compound eye allow easy access for producing a fragment corresponding to the human Aβ
phenotypic characterization. The fly’s behavior ranges from peptide [32]. Interestingly, this fragment is also able to
simple avoidance to learning and memory [23]. Due to its aggregate and induces age-dependent behavioral deficits
long history as an animal model in research, a wide variety and neurodegeneration [32].
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Table 1 Advantages and disadvantages of using Drosophila as a model organism for neurodegenerative diseases
like AD
Advantage Disadvantage
No ethical Brain anatomy, cardiovascular system and
problems/no restrictions according to animal protection laws respiration systems differs substantially from humans
Easy and cheap to maintain No easy measure of complex behavior
in large quantities, time and cost effective handling
Genetic manipulation is fast Only basic measures of cognitive decline
and inexpensive (3 month, < $ 500 per transgene)
Plethora of available Sometimes poor
resources/stocks (e.g. genome-wide RNAi-library) conservation of proteins/protein function
Short generation time (~10 days), Maintenance as living cultures only,
short life span (2–3 month)- > easy to use for screens no permanent conservation (e.g. frozen stocks) possible
Fully sequenced and annotated genome Less complex and adaptive
immune system as in vertebrates
Good conservation of basic Effects of drugs on the organism might differ
signaling pathways and cellular processes in general strongly (e.g. conversion of pro-toxins to toxins in liver)
Low redundancy/reduced
number of paralogous genes compared to vertebrates
Probably best
analyzed/understood multi- cellular organism
More complex
organism compare to C. elegans and yeast
Balancer chromosomes allow
the maintenance of mutations/trangenes without genotyping

In addition to endogenous Aβ production, transgenic by amyloidogenic processing of APP, influences that might
flies have been generated to study human Aβ42-induced result from APP processing are avoided. These flies have
toxicity and neurodegeneration [34-37]. Greeve and co- the major advantage of a direct assessment of Aβ toxicity.
workers generated a triple transgenic fly expressing human Neuronal expression of Aβ42 caused neurotoxicity,
APP (hAPP), human β-secretase (hBACE) and Drosophila locomotion defects and reduced lifespan. Moreover,
γ-secretase presenilin (dPsn) with point mutations corre- intra- and extracellular accumulation of Aβ42 peptides
sponding to familial AD mutations N141I, L235P and was observed. Overexpression of Aβ42[E22G], known to
E280A [36,38]. These flies developed age-dependent increase the rate of Aβ42 aggregation [7], exacerbated the
neurodegenerative phenotypes such as photoreceptor observed phenotypes [34]. Extensive investigation of
cell loss, severe degeneration of their projecting axons molecular mechanisms leading to changes in synaptic
and early lethality. Co-expression of hAPP and hBACE transmission and protein composition at the presynaptic
favored the processing of a higher glycosylated species of active zone revealed that Aβ42 expression affected axonal
hAPP in Drosophila resulting in Aβ40 and Aβ42 peptide transport of mitochondria and resulted in depletion of
forming plaques in transgene expressing tissue. Plaque mitochondria from the presynaptic active zone [48]. Intra-
deposition precedes the onset of neurodegeneration and neural accumulation of Aβ42 was shown to reduce synaptic
coexpression of mutant dPsn results in acceleration of vesicle release probability prior to bouton loss [49]. Patch
photoreceptor degeneration [36]. The described triple clamp analysis revealed a depression of cholinergic synapses
transgenic model clearly demonstrates the similarities upon Aβ42 expression. Moreover, expression of a familial
between the biochemical pathways induced by Aβ42 AD-linked mutant variant Aβ[E22G] caused an increased
deposition in flies and humans. aggregation of the Aβ42 peptide [50].
A more direct approach to investigate Aβ42-induced Finelli and co-workers established fly lines expressing
toxicity was used by Crowther and co-workers [34]. They fully processed, secreted Aβ peptides [35]. The generated
fused Aβ40/42 peptides to the signal peptide of endogen- transgenes allowed in-depth analysis of Aβ accumulation
ous Drosophila necrotic gene sequence ensuring secretion as overexpression of human Aβ40 and Aβ42 peptides can
[34]. Using the UAS/Gal4 inducible gene expression system be induced in a variety of cell types including neuronal
(Figure 1), the authors generated transgenic flies allowing cells. Both peptides accumulated in the fly brain but only
the spatiotemporal expression of Aβ40 and Aβ42. As the Aβ42 formed deposits [51]. Consequently, only Aβ42
expressed Aβ40/42 correspond to the peptides generated expressing flies show age-dependent and dose-dependent
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Figure 1 Genetic tools in Drosophila. In Drosophila the UAS/Gal4 expression system has been used extensively to express endogenous and
exogenous sequences in the tissue of interest [39]. This is implemented using two different lines. The so-called driver line contains a Gal4 coding
sequence inserted downstream of a promoter of an endogenous Drosophila gene. Gal4 is a transcription factor originating from Saccharomyces
cerevisiae [40]. It specifically binds to promoter elements termed upstream activating sequence (UAS), thus activating expression of the downstream
target sequence [40,41]. A collection of Gal4 driver lines which display a great variety of Gal4 expression in numerous tissues and organs is available to
the public [42]. Frequently used are the glass multimer reporter (GMR) driver inducing retinal expression [43] and the elav driver inducing pan-neuronal
expression [44]. After crossbreeding both, the Gal4 driver and the UAS line, the UAS target sequences will be expressed in a spatiotemporal manner
(depending on the Gal4 driver used). EP-elements are randomly inserted in the fly genome and contain UAS sites. Depending on the orientation
EP-elements might facilitate activation (same orientation) or inactivation (reverse orientation) of neighboring genes in a Gal4-dependent manner. There
are various collections of EP strains available allowing misexpression of a large number of fly genes [45,46]. So-called RNAi lines express short inverted
repeat sequences under UAS control. The sequence of the inverted repeat corresponds to an endogenous gene. Gal4-dependent expression of the
inverted repeat results in the formation short hairpin RNAs (shRNAs). The presence of shRNAs initiates a series of cellular mechanisms eventually
resulting in silencing of the corresponding endogenous gene by RNA interference (RNAi) [47].

neurodegeneration. In these flies, short-term memory was [52]. Positive effects of the mentioned drugs were also
impaired, obvious locomotor deficits appeared in aged evident in double transgenic AD mice overexpressing
flies and survival was reduced [37]. two mutated AD-linked transgenes (APPswe/PSEN1dE9)
As memory loss is a well-known feature of AD in [52,53]. Thus, results from invertebrate models systems
humans, memory assessment is widely used as an adequate might be well transferred to higher organisms.
tool to identify factors involved in Aβ42 pathomechanisms. Accumulating evidence suggests that impairment of metal
Recently, excess epidermal growth factor receptor (EGFR) homoeostasis is an important factor in AD pathogenesis.
was shown to enhance short-term memory loss in flies Levels of redox active metal ions such as copper, zinc and
concomitantly expressing Aβ42. The detrimental effect of iron are elevated in amyloid plaques of AD patients
EGFR overexpression on Aβ42-induced memory loss was [54]. Furthermore, it is known that presence of metals can
verified by the application of known EGFR inhibitors, e.g. promote Aβ aggregate formation in vitro and chelating
gefitinib and erlotinib. Both drugs are normally used in agents are able to dissolve Aβ plaques in post mortem AD
clinical cancer therapy, but were able to prevent Aβ42- brains [55,56].
induced memory loss in flies. Interestingly, also memantine, Drosophila models for AD proved to be a useful tool
a drug that is already used to treat dementia in AD patients, to investigate the influence of different metal ions on
prevented memory loss induced by Aβ42 expression in flies Aβ-induced neurodegeneration [57-61]. By feeding Aβ42
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expressing flies with copper or zinc supplemented food the expressing flies and simultaneously reduced the oxidative
Aβ42-induced phenotypes such as REP decreased survival damage in fly brains [61]. Thus, sequestration of free
and locomotor defects were enhanced. In contrast, food radicals by ferroxidase activity might be a beneficial
supplemented with metal-chelating substances suppressed mechanism protecting from oxidative stress originating
these phenotypes [57]. Genetic manipulation of metal from the redox potential of Aβ peptides in the Drosophila
homeostasis further underlined the role of zinc and copper model for Aβ42-induced toxicity [61].
levels in Aβ42-induced toxicity [57-59]. For example, Further adding to the topic of metal ions interacting
overexpression of MTF-1, a highly conserved transcription with Aβ peptides is a study about intrinsic toxicity of
factor inducing expression of several metal ion scavenger aluminum [62]. Typical neurodegenerative phenotypes
proteins, was shown to effectively protect from detrimental like reduced lifespan, locomotor deficits, olfactory learning
effects of Aß42 in flies [57]. Furthermore, genetic inhibition abnormalities and vacuolization of the brain were observed
of two copper-importers (Ctr1C and Ctr1B) ameliorated after feeding Drosophila with excess aluminum [62].
Aß42-induced neurodegenerative phenotypes while lower- Aluminum overload was shown to increase iron levels
ing copper load in the fly brain [58]. A study focusing on while simultaneously generating ROS. However, no direct
zinc as another redox active metal and its modulation link could be established between both processes [62].
of Aβ42-induced phenotypes basically showed the same Interestingly, expression of Aβ peptides or Tau did not
[59]. Genetic downregulation of the expression of the zinc modulate the Al-induced neurotoxicity [62]. This study
importer dZip1 consistently suppressed Aβ42-induced indicates that heavy metal ions can exert neurotoxic
brain vacuolization, locomotor defects and reduced life- effects per se and it remains to be elucidated if these
span, while overexpression had the opposite effect [59]. mechanisms are the cause or consequence in the interplay
Furthermore, the authors were able to show an effect between redox reactive metal ions, ROS generation and
of zinc deposition on the accumulation of Aβ fibrils in Aβ peptides.
Drosophila brains and a beneficial effect of dZip1 Apart from Aβ42 deposits, AD in humans is character-
knockdown on Aβ-induced early memory loss [59]. ized by intracellular neurofibrillary tangles composed
While findings about the detrimental effects of metal of hyperphosphorylated Tau proteins. As the functional
ion-Aβ complexes find a growing consent, not much is interactions between both AD lesions remain unclear, fly
known about the specific mechanisms of metal ions in AD. lines expressing Aβ42 were investigated for the formation
The study of Liu et al. took a closer look on the biophysical of fibrillary structures with fly endogenous Tau protein.
particularities of the interaction between iron and Aβ However, fibrillary structures composed of hyperpho-
peptides [60]. First, a connection between the presence sphorylated Tau could not be detected in Aβ42-expressing
of iron and modulation of Aβ42-induced toxicity was flies using biochemical or histological methods [51].
observed. Manipulation of the expression of iron-binding
proteins like ferritin and feeding of iron-specific chelating Drosophila models for Tau toxicity
agents altered Aβ42-induced toxicity [60]. Surprisingly, Insoluble aggregates of the MT-associated protein Tau
knockdown of ferritin did not reduce Aβ accumulation are a common feature of so-called tauopathies like fronto-
but efficiently suppressed Aβ42-induced toxicity [60]. temporal dementia with parkinsonism linked to chromo-
Instead, biophysical techniques revealed that the presence some 17 (FTDP-17), progressive supranuclear palsy
of iron during Aβ42 aggregation altered the structure of and Pick’s disease and others [63]. Central feature of
Aβ fibrils delaying the formation of mature aggregates tauopathies is the presence of paired helical filaments,
[60]. Cytotoxicity assays using human neuroblastoma which assemble into intracellular neurofibrillary tangles in
SH-SY5Y cells indicated that the presence of iron during affected tissues [64]. Several disease-linked mutations in
aggregate formation was contributing to Aβ toxicity rather the Tau gene affect correct splicing of its MT binding sites,
than addition of iron after aggregate formation [60]. Thus, thus enhancing abnormal phosphorylation and detachment
the authors conclude that modulation of the kinetics of Aβ of the protein. Both steps are believed to be crucial in
aggregate formation by iron is important for the toxicity of the process of forming paired helical filaments and higher
Aβ42 peptides [60]. order neurofibrillary tangles [65,66].
Besides the potential function of metal ions to act as Overexpression of wild-type or mutant human Tau in
seeds for Aβ accumulation, they might also play a role the Drosophila nervous system caused vacuolization in the
in the production of reactive oxygen species (ROS) via brain accompanied by pathologic phosphorylation status of
Fenton-like reactions. An unbiased screen identified many Tau, although large filamentous aggregates were absent
modifiers of Aβ42-induced toxicity that were implicated [64]. Nevertheless, immunostaining with antibodies detect-
in redox regulation [61]. Overexpression of two subunits of ing abnormal confirmation of Tau revealed a close associ-
ferritin, a highly conserved protein with a strong antioxi- ation between areas of degeneration and abnormal Tau in
dant potential, efficiently prolonged the lifespan of Aβ42 flies. Moreover, the abundance of vacuolar lesions in the fly
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brain was first observed in Tau expressing tissue. In Chatterjee et al. created fly lines expressing phosphoryl-
addition, neurodegeneration progressed with fly age and ation-resistant Tau variants by exchanging two (TauS2A) or
eventually resulted in early mortality. Furthermore, severity eleven (TauS11A) putative serine-threonine phosphorylation
of phenotypes was enhanced by increasing Tau dosage sites with neutral alanine. These mutations prevented phos-
or introducing mutant Tau isoforms, such as the V337M phorylation by protease activated receptor 1 (PAR-1) and
and R406W mutations associated with FTDP-17 [64]. In GSK3β, respectively [67]. This allowed a thorough investi-
addition, targeted expression of either wild-type or mutant gation of several Tau kinases in disease-related processes
Tau in the retina caused alterations in external eye such as site-specific phosphorylation and changes in
structures, characterized by size reduction and rough MT binding properties of Tau [67]. Interestingly, REP
appearance. The so-called rough eye phenotype (REP) enhancement induced by overexpression of GSK3β was
correlates with the loss of retinal cells including photore- less pronounced in the TauS2A expressing fly compared to
ceptors [63,64,67,68]. Detailed analysis revealed that Tau the wild-type Tau expressing fly although immunoblotting
overexpression caused degeneration of photoreceptor axons, using phosphorylation site-specific Tau antibodies showed
evident by the appearance of vacuoles in the medulla, a higher degree of Tau phosphorylation. In contrast,
the projection target of photoreceptor axons [63]. Such TauS11A was resistant to GSK3β phosphorylation although
REPs are frequently used to screen for genetic interactions GSK3β overexpression enhanced the TauS2A-induced REP
(see Table 2). In such an approach the fly ortholog of severity. Furthermore, neither Tau aggregation nor MT
glycogen synthase kinase 3β (GSK3β) was identified to binding properties consistently correlated with REP [67].
interfere with Tau-induced toxicity. Interestingly, the These results uncouple Tau toxicity from sole phosphoryl-
Tau-induced REP was suppressed in a GSK3β-deficient ation and indicate Tau toxicity is partially independent of
background and enhanced by GSK3β overexpression its phosphorylation state.
[68]. Detailed analysis showed that overexpression of In addition, Iijima-Ando et al. generated another
GSK3β strongly increased pathogenic phosphorylation phosphorylation-resistant Tau variant TauS262A [73]. Retinal
of Tau [68,71]. coexpression of wild-type human Tau and DNA damage-
In order to investigate the role of Tau phosphorylation activated checkpoint kinase 2 (Chk2) resulted in enhance-
and toxicity in more detail, several Tau variants with ment of the REP. In contrast, coexpression of Chk2 and
altered phosphorylation sites were generated [67,73,74]. TauS262A had no effect on eye surface integrity [73].

Table 2 Overview of performed large-scale screens for modifiers of toxicity induced by expression of AD-linked genes
in Drosophila melanogaster
Transgene causing a REP Screened library Results Reference
hTau[V337M] 2,276 EP strains • Kinases, phosphatases Shulman & Feany [69]
(CDK5, GSK3β, PAR1)
• Apoptosis
• Novel: Ataxin 2, Fmr1
hTau[V337M] 1,250 P-element strains • Cytoskeletal components Blard et al. [70]
• Molecular chaperones
• Chromatin remodelling
hTau[WT] 920 P-lethal strains • Kinases, Phosphatases Ambegaokar et al. [71]
895 EY strains • Autophagy/lysosomal
• RNA processing
• Chromatin regulation
• Cytoskeletal
GMR > Aβ42 1,963 EP strains • Secretory pathway Cao et al. [72]
• Cholesterol homeostasis
• Chromatin regulation
Pan neural Arctic 3,000 de novo • Fenton chemistry and oxidative Rival et al. [61]
Aβ42 life span reduction insertions of transposable elements stress are involved in AD pathology
• Ferritin expression
protects from β-amyloid toxicity
The table lists only screens in which the fly was used as primary screening tool. Not listed are screens using other sources to gain candidates, later confirmed
in flies.
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To determine the contribution of specific phosphoryl- reading frame, Gal4 induces either ectopic overexpression
ation sites to Tau toxicity, Steinhilb et al. designed novel or inactivation of the gene by RNA interference (RNAi)
Tau transgenes [74]. By replacing serines of several [45]. After comprehensive validation of identified candi-
disease-associated phosphorylation sites with alanine dates they were functionally classified. The largest group
they created a phosphorylation-resistant variant (TauAP) of modifiers were kinases and phosphatases. Among these
and by replacing serines with glutamines they mimicked a kinases were Drosophila orthologs of known Tau kinases
hyperphosphorylated state of Tau (TauE14). The conse- such as cyclin-dependent kinase 5 (CDK5) and GSK3β.
quences are amelioration of Tau toxicity in flies expressing Accordingly, these results confirmed the reliability of
phospho-deficient Tau variant TauAP and exacerbation of the screening approach and emphasizes the critical role of
Tau toxicity in flies expressing the phospho-mimetic Tau Tau phosphorylation for toxicity [69].
variant TauE14 [74]. However, mutation of individual serines Using the same transgenic fly line expressing human
of the respective phosphorylation sites did not result in a Tau [V337M], Blard et al. screened a different collection
clear modulation of Tau toxicity indicating that multiple of 1,250 EP-element containing fly lines [70]. According
sites work in concert to confer to Tau toxicity [75]. to the differences in fly lines and the low percentage of
Folwell and co-workers analyzed concomitant expression whole genome coverage, there was little overlap between
of Aβ42 and Tau in flies. In these flies, Aβ42 expression identified modifiers from this screen compared to the
exacerbated Tau-induced neuronal dysfunction, axonal screen by Shulman and Feany. Blard et al. identified
transport deficits and decreased survival [76]. The com- several components of the cytoskeleton as modifiers of
binatorial expression of both pathological proteins Aβ42 Tau-induced REP. In addition, the Tau-induced disruption
and Tau in Drosophila seems to be a promising approach of the MT network at nerve terminals was identified as
to investigate the synergistic effects at the level of genetic key event leading to Tau-induced neurodegeneration [70].
interactions. The most recent large-scale screen for modifiers of Tau
toxicity was performed by Ambegaokar et al. [71]. In their
Large-scale screens in Drosophila screen, the authors used a fly line expressing wild-type
Low demand on care and easiness of handling predestine human Tau in the fly eye. This fly line also exhibited an
the fly to high-throughput screens in vivo. Adding to these intermediate REP, which was suitable to identify both
advantages is the extraordinary large pool of available enhancer and suppressors. The authors screened two
genetic instruments paired with simplicity of the genomic independent collections of fly lines. The first contains
structure facilitating subsequent in-depth analysis. roughly 1,000 lethal loss-of-function alleles caused by
Up to now unbiased screens in Drosophila were per- P-element insertion in essential genes. The second col-
formed utilizing the above-described tools and provided lection contained 900 lines with random insertions of
valuable insights into AD pathomechanisms (see Table 2) EY-elements. These EY-elements are very similar to
[69-72]. REPs induced by expression of toxic gene products EP-elements and also contain UAS sites. Once Gal4
in the Drosophila compound eye represent an easy to score is present, this can result in overexpression or RNAi-
read-out for genetic modifier screens. The fly eye is a mediated silencing of genes in close vicinity to the
neuronal structure and REPs are highly sensitive to genetic insertion site of the element (Figure 1). In their screen,
modification. Changes in REP severity usually coincide Ambegaokar and co-workers identified known interac-
with changes in photoreceptor degeneration, thus changes tors of Tau toxicity such as the Drosophila ortholog of
in neuronal decline can be investigated by light microscopy GSK3β. This can be regarded as validation of the screen
(Figure 2). and suggests that identified modifiers could be relevant
Shulman and Feany conducted the first large-scale to disease. Comprehensive analysis of identified modifiers
screen in Drosophila for genetic modifiers of toxicity using computational network approach revealed a
induced by expression of human Tau [69]. In their screen, broad range of functional classes including kinases,
the authors used the fact that eye-specific expression cytoskeletal components as expected but also mechanisms
of a FTLD-linked Tau variant (Tau[V337M]) induced a not yet associated to Tau toxicity such as RNA metabolism
moderate REP. To facilitate identification of enhancers or chromatin interaction [71]. Furthermore, the authors
and suppressors, flies with the Tau-dependent REP were found that differences in Tau phosphorylation did not
crossbred with a collection of 2,276 enhancer promoter correlate with changes in Tau toxicity [71].
(EP) insertion-carrying flies. These files contain random Only few large-scale screens have been published
insertions of EP-elements, which can be used to misexpress identifying genetic modifiers of Aβ42-induced toxicity
endogenous fly genes (Figure 1) [45]. EP-elements contain (see Table 2 and [61,72]). Cao et al. screened a collection of
UAS sites allowing the Gal4-induced transcription of open EP-element carrying fly lines for modification of Aβ42-
reading frames in the vicinity of insertion. Depending on induced REP in Drosophila [72]. Modifiers identified in this
the orientation of the EP-element in relation to the open screen comprise loss-of-function alleles widely involved in
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Figure 2 Exemplified rough eye phenotypes (REP) used as readout for modifier screens. Scanning electron micrographs (top) of fly eyes
are shown. The Drosophila compound eye consists of a stereotypic array of about 800 omatidia (left). These hexagonal structures are highly
ordered and display regular spacing of hairs called interomatidial bristles (inset). Expression of disease-linked proteins/peptides in the eye can
cause a REP (middle). The rough appearance of the eye can be caused by loss of interomatidial bristles, fusion of omatidia, necrotic tissue, dints
in the retina and is often accompanied by loss of pigmentation and reduced eye size. An enhancement in severity (left) is easily observable by
more pronounced REP characteristics. Usually, such REPs are sensitive towards genetic interactions, causing either a suppression (left) or an
enhancement (right), changing the overall eye appearance towards a more wild-type like appearance (suppression) or by increasing the rough
appearance of the eye (enhancement), respectively. Exemplary light micrographs show REPs induced by expression of either Tau[R406W] (middle)
or Aβ42 (bottom). These REPs are sensitive towards genetic modification like suppression (left) and enhancement (right) and can be/have been
used for screening approaches.

cell compartment trafficking pathways leading to the con- known open reading frames of the Drosophila genome.
clusion that proper function of endocytosis and vesicular Using an inducible short hairpin RNA (shRNA) expressing
trafficking is critical to protect the cell from Aβ42-induced fly line, the RNAi effect can be activated in a spatio-
toxicity. In addition, a reasonable number of candidate temporal manner (Figure 1). Recently, an in vivo RNAi
genes involved in secretory pathways were identified. library was generated utilizing the UAS/Gal4 system to
Thus, the authors argue that proteolytic degradation of Aβ control shRNA expression [47].
peptides during translocation by the secretory pathways The RNAi library has been extensively used for genome-
might be a crucial pathomechanism in AD [72]. On the wide, large-scale screens to identify genetic modifiers of
other hand, Rival and co-workers convincingly showed that basic cellular mechanisms [77-79]. However, published
Fenton chemistry and oxidative stress contribute to the data regarding the above-described Aβ42 toxicity models are
toxicity of β-amyloid peptides in flies [61]. surprisingly scarce [72,80]. Nevertheless, this approach has
The combination of the Aβ42-induced REP with the been used to find genetic modifiers of Ataxin-3-derived
utilization of RNAi allows for an unbiased screen targeting polyglutamine-induced toxicity [81]. The analysis yielded a
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large number of genetic modifiers that imply involvement AD. In vitro assays proved the activity of several engineered
of multiple processes in polyglutamine toxicity. β-secretase inhibitors but many failed in cellular assays
To aid the understanding of mechanisms leading to [85,86]. However, in vivo the endosomal localization of
AD, we performed a genome-wide screen for modifiers β-secretase is essential for activity. Coupling of a sterol
of Aβ42-induced neurodegeneration [82]. By combining moiety to the inhibitor resulted in successful delivery to
eye-specific RNAi-mediated knockdown of single Dros- the endosomal membrane and efficient inhibition of
ophila genes and concomitant Aβ42 expression, genetic β-secretase cleavage of APP in several cell lines [84].
interactors modulating Aβ42-induced REP were identified Furthermore, inhibition of β-secretase activity by the
and were assigned to cellular pathways contributing to sterol-coupled inhibitor was shown to be efficient in vivo
Aβ42 toxicity. To prove adaptability of the performed using the triple transgenic fly line expressing hAPP,
screen, we tested RNAi lines targeting corresponding hBACE and dPsn created by Greeve et al. [36]. Transgenic
Drosophila orthologs of known susceptibility genes identi- larvae fed with the membrane-tethered steady-state in-
fied by genome-wide association studies (GWAS) for their hibitor showed increased hatching rates compared to
ability to modulate the Aβ42-induced REP. Preliminary transgenic larvae fed with soluble inhibitor [84]. Thus,
results indicate low conformity between the effects of flies expressing disease-related transgenes might be very
RNAi-mediated knockdown of susceptibility genes and useful to prove hypotheses in vivo in a fast, effective and
enhancement or suppression of Aβ42-induced REP (un- economic manner.
published results). One way to explain this might be the Despite the efforts of countless scientists worldwide to
redundancy of affected pathways. Another possibility clarify the mechanisms underlying the most prevalent
might be low penetrance of the RNAi effect, although form of dementia, it is still not possible to cure AD. Until
the majority of the RNAi library was tested for effective now therapies for AD have included only symptomatic
silencing of targeted genes [47]. Still, AD is not a mono- treatment and there is not even any effective medication
genic disease and application of GWAS to identify human to stop disease progression. The mere number of hypoth-
risk factors failed to find new major genes relevant to all eses intending to explain the pathogenesis of AD hints at
AD patients [83]. In addition, we conducted a very similar the general challenge this disease poses to modern
screen to identify modifiers of Tau[R406W]-induced science. The challenge now is to elucidate the contribution
neurodegeneration. To our surprise, in this screen we only of AD-associated pathways with known effects to Aβ42-
identified a very small amount of modifiers (less than 100 induced neurodegeneration and to differentiate the path-
out of roughly 8,000 screened RNAi lines modified the ways modifying general neurodegenerative mechanisms
Tau[R406W]-induced REP). Among the few candidates from the ones that are unique to AD and thus provide
were members of the dynein/dynactin complex. As silen- a target for drug development.
cing members of the dynein/dynactin complex enhanced
the Tau[R406W]-induced toxicity, an impaired retrograde
axonal transport seems to contribute to Tau[R406W]- Ethical approval
induced toxicity (to be published elsewhere). Experimental research reported here was performed using
insects (Drosophilae). Such research is exempt from
Perspectives and conclusion regulations pertaining to ethical approvals and/or animal
Drosophila melanogaster is a useful in vivo tool to analyze protection laws.
pathomechanisms in AD. For example, aggregation of
Aβ42 can be easily determined in flies. Thus, large collec- Abbreviations
Aβ: Amyloid-β; AD: Alzheimer’s disease; APP: Amyloid precursor
tions of small compounds can be screened for their protein; BACE: β-site APP-cleaving enzyme; dAPPl: APP-like,
potency to inhibit Aβ peptide aggregation [80]. Recently, a Drosophila melanogaster ortholog of APP; dPsn: Drosophila
compound (D737) was identified that effectively inhibited melanogaster ortholog of presenilin; EGFR: Epidermal growth
factor receptor; EP: Enhancer-promoter; FTDP-17: Frontotemporal
fibril formation in vitro. Administration of this compound dementia with parkinsonism linked to chromosome 17; GSK3β:
to flies prevented early death usually observed after Aβ42 Glycogen synthase kinase 3β; GWAS: Genome-wide association studies;
expression [80]. Such in vivo approaches might help in MT: Microtubuli; PAR1: Protease activated receptor 1; REP: Rough eye
phenotype; RNAi: RNA interference; shRNA: short hairpin RNA;
drug development not only in case of AD, but also in the UAS: Upstream activating sequence.
context of other (neurodegenerative) diseases.
Furthermore, transgenic fly lines can be used to prove
Competing interests
efficiency of β-secretase steady-state inhibitors [84]. The authors declare that they have no competing interests.
β-Secretase activity is the rate-limiting step during amyloi-
dogenic processing leading to the generation of pathogenic
Authors’ contributions
Aβ peptides. Thus, β-secretase activity is a preferred target KP and AV wrote the manuscript. JBS wrote and drafted the manuscript. All
for the development of pharmacological therapies against authors read and approved the final manuscript.
Prüßing et al. Molecular Neurodegeneration 2013, 8:35 Page 10 of 11
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Author details 21. Adams MD, Celniker SE, Holt RA, Evans CA, Gocayne JD, Amanatides PG,
1
Department of Neurology, University Medical Center, RWTH Aachen, Scherer SE, Li PW, Hoskins RA, Galle RF, et al: The genome sequence of
Pauwelsstrasse 30, D-52074 Aachen, Germany. 2Jülich-Aachen Research Drosophila melanogaster. Science 2000, 287:2185–2195.
Alliance (JARA) Brain, Aachen, 52074, Germany. 3EURON - European Graduate 22. Nichols CD: Drosophila melanogaster neurobiology, neuropharmacology,
School of Neuroscience, Aachen, Germany. and how the fly can inform central nervous system drug discovery.
Pharmacol Ther 2006, 112:677–700.
Received: 21 June 2013 Accepted: 13 November 2013 23. McGuire SE, Deshazer M, Davis RL: Thirty years of olfactory learning and
Published: 22 November 2013 memory research in Drosophila melanogaster. Prog Neurobiol 2005,
76:328–347.
24. Greenspan RJ: Fly Pushing: The Theory and Practice of Drosophila Genetics.
References New Jersey: Cold Spring Harbour Laboratory Press; 2004.
1. Ferri CP, Prince M, Brayne C, Brodaty H, Fratiglioni L, Ganguli M, Hall K,
25. Lenz S, Karsten P, Schulz JB, Voigt A: Drosophila as a screening tool to
Hasegawa K, Hendrie H, Huang Y, et al: Global prevalence of dementia: a
study human neurodegenerative diseases. J Neurochem 2013, 127:453–60.
Delphi consensus study. Lancet 2005, 366:2112–2117.
26. Bonner JM, Boulianne GL: Drosophila as a model to study age-related
2. Chow VW, Mattson MP, Wong PC, Gleichmann M: An overview of APP
neurodegenerative disorders: Alzheimer’s disease. Exp Gerontol 2011,
processing enzymes and products. Neuromolecular Med 2010, 12:1–12.
46:335–339.
3. Burdick D, Soreghan B, Kwon M, Kosmoski J, Knauer M, Henschen A, Yates J,
27. Cowan CM, Shepherd D, Mudher A: Insights from Drosophila models of
Cotman C, Glabe C: Assembly and aggregation properties of synthetic
Alzheimer’s disease. Biochem Soc Trans 2010, 38:988–992.
Alzheimer’s A4/beta amyloid peptide analogs. J Biol Chem 1992,
28. Rubin GM, Yandell MD, Wortman JR, Gabor Miklos GL, Nelson CR, Hariharan
267:546–554.
IK, Fortini ME, Li PW, Apweiler R, Fleischmann W, et al: Comparative
4. Iqbal K, Liu F, Gong CX, Grundke-Iqbal I: Tau in Alzheimer disease and re-
genomics of the eukaryotes. Science 2000, 287:2204–2215.
lated tauopathies. Curr Alzheimer Res 2010, 7:656–664.
29. Fortini ME, Skupski MP, Boguski MS, Hariharan IK: A survey of human
5. Hardy JA, Higgins GA: Alzheimer’s disease: the amyloid cascade
disease gene counterparts in the Drosophila genome. J Cell Biol 2000,
hypothesis. Science 1992, 256:184–185.
150:F23–30.
6. Bentahir M, Nyabi O, Verhamme J, Tolia A, Horre K, Wiltfang J, Esselmann H,
30. Luo L, Tully T, White K: Human amyloid precursor protein ameliorates
de Strooper B: Presenilin clinical mutations can affect gamma-secretase
behavioral deficit of flies deleted for Appl gene. Neuron 1992, 9:595–605.
activity by different mechanisms. J Neurochem 2006, 96:732–742.
7. Nilsberth C, Westlind-Danielsson A, Eckman CB, Condron MM, Axelman K, 31. Periz G, Fortini ME: Functional reconstitution of gamma-secretase through
Forsell C, Stenh C, Luthman J, Teplow DB, Younkin SG, et al: The ‘Arctic’ coordinated expression of presenilin, nicastrin, Aph-1, and Pen-2.
APP mutation (E693G) causes Alzheimer’s disease by enhanced Abeta J Neurosci Res 2004, 77:309–322.
protofibril formation. Nat Neurosci 2001, 4:887–893. 32. Carmine-Simmen K, Proctor T, Tschape J, Poeck B, Triphan T, Strauss R,
8. Suzuki N, Cheung TT, Cai XD, Odaka A, Otvos L Jr, Eckman C, Golde TE, Kretzschmar D: Neurotoxic effects induced by the Drosophila amyloid-
Younkin SG: An increased percentage of long amyloid beta protein beta peptide suggest a conserved toxic function. Neurobiol Dis 2009,
secreted by familial amyloid beta protein precursor (beta APP717) 33:274–281.
mutants. Science 1994, 264:1336–1340. 33. Yagi Y, Tomita S, Nakamura M, Suzuki T: Overexpression of human
9. Spillantini MG, Murrell JR, Goedert M, Farlow MR, Klug A, Ghetti B: Mutation amyloid precursor protein in Drosophila. Mol Cell Biol Res Commun 2000,
in the tau gene in familial multiple system tauopathy with presenile 4:43–49.
dementia. Proc Natl Acad Sci USA 1998, 95:7737–7741. 34. Crowther DC, Kinghorn KJ, Miranda E, Page R, Curry JA, Duthie FA, Gubb
10. Hutton M, Lendon CL, Rizzu P, Baker M, Froelich S, Houlden H, Pickering-Brown DC, Lomas DA: Intraneuronal Abeta, non-amyloid aggregates and
S, Chakraverty S, Isaacs A, Grover A, et al: Association of missense and neurodegeneration in a Drosophila model of Alzheimer’s disease.
5′-splice-site mutations in tau with the inherited dementia FTDP-17. Neuroscience 2005, 132:123–135.
Nature 1998, 393:702–705. 35. Finelli A, Kelkar A, Song HJ, Yang H, Konsolaki M: A model for studying
11. Oddo S, Caccamo A, Cheng D, Jouleh B, Torp R, LaFerla FM: Genetically Alzheimer’s Abeta42-induced toxicity in Drosophila melanogaster.
augmenting tau levels does not modulate the onset or progression of Mol Cell Neurosci 2004, 26:365–375.
Abeta pathology in transgenic mice. J Neurochem 2007, 102:1053–1063. 36. Greeve I, Kretzschmar D, Tschape JA, Beyn A, Brellinger C, Schweizer M,
12. Roberson ED, Scearce-Levie K, Palop JJ, Yan F, Cheng IH, Wu T, Gerstein H, Nitsch RM, Reifegerste R: Age-dependent neurodegeneration and
Yu GQ, Mucke L: Reducing endogenous tau ameliorates amyloid Alzheimer-amyloid plaque formation in transgenic Drosophila. J Neurosci
beta-induced deficits in an Alzheimer’s disease mouse model. 2004, 24:3899–3906.
Science 2007, 316:750–754. 37. Iijima K, Chiang HC, Hearn SA, Hakker I, Gatt A, Shenton C, Granger L, Leung
13. Ittner LM, Ke YD, Delerue F, Bi M, Gladbach A, van Eersel J, Wolfing H, A, Iijima-Ando K, Zhong Y: Abeta42 mutants with different aggregation
Chieng BC, Christie MJ, Napier IA, et al: Dendritic function of tau mediates profiles induce distinct pathologies in Drosophila. PLoS One 2008,
amyloid-beta toxicity in Alzheimer’s disease mouse models. Cell 2010, 3:e1703.
142:387–397. 38. Ye Y, Fortini ME: Apoptotic activities of wild-type and Alzheimer’s
14. Winderickx J, Delay C, de Vos A, Klinger H, Pellens K, Vanhelmont T, van disease-related mutant presenilins in Drosophila melanogaster.
Leuven F, Zabrocki P: Protein folding diseases and neurodegeneration: J Cell Biol 1999, 146:1351–1364.
lessons learned from yeast. Biochim Biophys Acta 2008, 1783:1381–1395. 39. Phelps CB, Brand AH: Ectopic gene expression in Drosophila using GAL4
15. Teschendorf D, Link CD: What have worm models told us about the system. Methods 1998, 14:367–379.
mechanisms of neuronal dysfunction in human neurodegenerative 40. Lohr D, Venkov P, Zlatanova J: Transcriptional regulation in the yeast GAL
diseases? Mol Neurodegener 2009, 4:38. gene family: a complex genetic network. FASEB J 1995, 9:777–787.
16. Gotz J, Chen F, Barmettler R, Nitsch RM: Tau filament formation in 41. Ptashne M: How eukaryotic transcriptional activators work. Nature 1988,
transgenic mice expressing P301L tau. J Biol Chem 2001, 276:529–534. 335:683–689.
17. Hsiao K, Chapman P, Nilsen S, Eckman C, Harigaya Y, Younkin S, Yang F, 42. Bloomington Stock Center. [http://flystocks.bio.indiana.edu/Browse/in/
Cole G: Correlative memory deficits, Abeta elevation, and amyloid misexpression-top.php]
plaques in transgenic mice. Science 1996, 274:99–102. 43. Moses K, Ellis MC, Rubin GM: The glass gene encodes a zinc-finger protein
18. Sturchler-Pierrat C, Abramowski D, Duke M, Wiederhold KH, Mistl C, required by Drosophila photoreceptor cells. Nature 1989, 340:531–536.
Rothacher S, Ledermann B, Burki K, Frey P, Paganetti PA, et al: Two amyloid 44. Brand AH, Perrimon N: Targeted gene expression as a means of altering
precursor protein transgenic mouse models with Alzheimer disease-like cell fates and generating dominant phenotypes. Development 1993,
pathology. Proc Natl Acad Sci USA 1997, 94:13287–13292. 118:401–415.
19. Mhatre SD, Paddock BE, Saunders AJ, Marenda DR: Invertebrate models of 45. Rorth P: A modular misexpression screen in Drosophila detecting
Alzheimer’s disease. J Alzheimers Dis 2013, 33:3–16. tissue-specific phenotypes. Proc Natl Acad Sci USA 1996, 93:12418–12422.
20. Morgan TH: Sex Limited Inheritance in Drosophila. Science 1910, 46. Szeged Drosophila Stock Center. [http://expbio.bio.u-szeged.hu/fly/modules.
32:120–122. php?name = Other_Stocks&op = OtherStocksList&stock_gr = 4]
Prüßing et al. Molecular Neurodegeneration 2013, 8:35 Page 11 of 11
http://www.molecularneurodegeneration.com/content/8/1/35

47. Dietzl G, Chen D, Schnorrer F, Su KC, Barinova Y, Fellner M, Gasser B, Kinsey 67. Chatterjee S, Sang TK, Lawless GM, Jackson GR: Dissociation of tau toxicity
K, Oppel S, Scheiblauer S, et al: A genome-wide transgenic RNAi library for and phosphorylation: role of GSK-3beta, MARK and Cdk5 in a Drosophila
conditional gene inactivation in Drosophila. Nature 2007, 448:151–156. model. Hum Mol Genet 2009, 18:164–177.
48. Zhao XL, Wang WA, Tan JX, Huang JK, Zhang X, Zhang BZ, Wang YH, 68. Jackson GR, Wiedau-Pazos M, Sang TK, Wagle N, Brown CA, Massachi S,
YangCheng HY, Zhu HL, Sun XJ, Huang FD: Expression of beta-amyloid Geschwind DH: Human wild-type tau interacts with wingless pathway
induced age-dependent presynaptic and axonal changes in Drosophila. components and produces neurofibrillary pathology in Drosophila.
J Neurosci: the official journal of the Society for Neuroscience 2010, Neuron 2002, 34:509–519.
30:1512–1522. 69. Shulman JM, Feany MB: Genetic modifiers of tauopathy in Drosophila.
49. Huang JK, Ma PL, Ji SY, Zhao XL, Tan JX, Sun XJ, Huang FD: Age-dependent Genetics 2003, 165:1233–1242.
alterations in the presynaptic active zone in a Drosophila model of 70. Blard O, Feuillette S, Bou J, Chaumette B, Frebourg T, Campion D, Lecourtois
Alzheimer’s disease. Neurobiol Dis 2013, 51:161–167. M: Cytoskeleton proteins are modulators of mutant tau-induced neuro-
50. Fang L, Duan J, Ran D, Fan Z, Yan Y, Huang N, Gu H, Zhu Y: Amyloid-beta degeneration in Drosophila. Hum Mol Genet 2007, 16:555–566.
depresses excitatory cholinergic synaptic transmission in Drosophila. 71. Ambegaokar SS, Jackson GR: Functional genomics screen and network
Neurosci Bull 2012, 28:585–594. analysis reveal novel modifiers of tauopathy dissociated from tau
51. Iijima K, Liu HP, Chiang AS, Hearn SA, Konsolaki M, Zhong Y: Dissecting the phosphorylation. Hum Mol Genet 2011, 20:4947–4977.
pathological effects of human Abeta40 and Abeta42 in Drosophila: a 72. Cao W, Song HJ, Gangi T, Kelkar A, Antani I, Garza D, Konsolaki M: Identification
potential model for Alzheimer’s disease. Proc Natl Acad Sci USA 2004, of novel genes that modify phenotypes induced by Alzheimer’s beta-amyloid
101:6623–6628. overexpression in Drosophila. Genetics 2008, 178:1457–1471.
52. Wang L, Chiang HC, Wu W, Liang B, Xie Z, Yao X, Ma W, Du S, Zhong Y: 73. Iijima-Ando K, Zhao L, Gatt A, Shenton C, Iijima K: A DNA damage-activated
Epidermal growth factor receptor is a preferred target for treating checkpoint kinase phosphorylates tau and enhances tau-induced
amyloid-beta-induced memory loss. Proc Natl Acad Sci USA 2012, neurodegeneration. Hum Mol Genet 2010, 19:1930–1938.
109:16743–16748. 74. Steinhilb ML, Dias-Santagata D, Mulkearns EE, Shulman JM, Biernat J,
53. Martinez-Coria H, Green KN, Billings LM, Kitazawa M, Albrecht M, Rammes G, Mandelkow EM, Feany MB: S/P and T/P phosphorylation is critical for tau
Parsons CG, Gupta S, Banerjee P, LaFerla FM: Memantine improves neurotoxicity in Drosophila. J Neurosci Res 2007, 85:1271–1278.
cognition and reduces Alzheimer’s-like neuropathology in transgenic 75. Steinhilb ML, Dias-Santagata D, Fulga TA, Felch DL, Feany MB: Tau phos-
mice. Am J Pathol 2010, 176:870–880. phorylation sites work in concert to promote neurotoxicity in vivo. Mol
54. Lovell MA, Robertson JD, Teesdale WJ, Campbell JL, Markesbery WR: Biol Cell 2007, 18:5060–5068.
Copper, iron and zinc in Alzheimer’s disease senile plaques. J Neurol Sci 76. Folwell J, Cowan CM, Ubhi KK, Shiabh H, Newman TA, Shepherd D, Mudher
1998, 158:47–52. A: Abeta exacerbates the neuronal dysfunction caused by human tau
55. Atwood CS, Moir RD, Huang X, Scarpa RC, Bacarra NM, Romano DM, expression in a Drosophila model of Alzheimer’s disease. Exp Neurol 2010,
Hartshorn MA, Tanzi RE, Bush AI: Dramatic aggregation of Alzheimer abeta 223:401–409.
by Cu(II) is induced by conditions representing physiological acidosis. 77. Enneking EM, Kudumala SR, Moreno E, Stephan R, Boerner J,
J Biol Chem 1998, 273:12817–12826. Godenschwege TA, Pielage J: Transsynaptic coordination of synaptic
56. Cherny RA, Legg JT, McLean CA, Fairlie DP, Huang X, Atwood CS, growth, function, and stability by the L1-Type CAM Neuroglian. PLoS Biol
Beyreuther K, Tanzi RE, Masters CL, Bush AI: Aqueous dissolution of 2013, 11:e1001537.
Alzheimer’s disease Abeta amyloid deposits by biometal depletion. 78. Read RD, Fenton TR, Gomez GG, Wykosky J, Vandenberg SR, Babic I,
J Biol Chem 1999, 274:23223–23228. Iwanami A, Yang H, Cavenee WK, Mischel PS, et al: A kinome-wide RNAi
57. Hua H, Munter L, Harmeier A, Georgiev O, Multhaup G, Schaffner W: screen in Drosophila Glia reveals that the RIO kinases mediate cell prolif-
Toxicity of Alzheimer’s disease-associated Abeta peptide is ameliorated eration and survival through TORC2-Akt signaling in glioblastoma. PLoS
in a Drosophila model by tight control of zinc and copper availability. Genet 2013, 9:e1003253.
Biol Chem 2011, 392:919–926. 79. Zhang J, Liu M, Su Y, Du J, Zhu AJ: A targeted in vivo RNAi screen reveals
58. Lang M, Fan Q, Wang L, Zheng Y, Xiao G, Wang X, Wang W, Zhong Y, Zhou deubiquitinases as new regulators of Notch signaling. G3 (Bethesda) 2012,
B: Inhibition of human high-affinity copper importer Ctr1 orthologous in 2:1563–1575.
the nervous system of Drosophila ameliorates Abeta42-induced 80. McKoy AF, Chen J, Schupbach T, Hecht MH: A novel inhibitor of amyloid beta
Alzheimer’s disease-like symptoms. Neurobiol Aging 2013, 34:2604–2612. (Abeta) peptide aggregation: from high throughput screening to efficacy in
59. Lang M, Wang L, Fan Q, Xiao G, Wang X, Zhong Y, Zhou B: Genetic an animal model of Alzheimer disease. J Biol Chem 2012, 287:38992–39000.
inhibition of solute-linked carrier 39 family transporter 1 ameliorates 81. Voßfeldt H, Butzlaff M, Prüßing K, Ni Charthaigh RA, Karsten P, Lankes A,
abeta pathology in a Drosophila model of Alzheimer’s disease. Hamm S, Simons M, Adryan B, Schulz JB, Voigt A: Large-scale screen for
PLoS Genet 2012, 8:e1002683. modifiers of ataxin-3-derived polyglutamine-induced toxicity in
Drosophila. PLoS One 2012, 7:e47452.
60. Liu B, Moloney A, Meehan S, Morris K, Thomas SE, Serpell LC, Hider R,
82. Prüßing K, Voigt A, Schulz JB: Genome-wide screen for modifiers of Ab42-
Marciniak SJ, Lomas DA, Crowther DC: Iron promotes the toxicity of
induced neurodegeneration in Drosophila. Wednesday 31 August 2011.
amyloid beta peptide by impeding its ordered aggregation. J Biol Chem
J Neurochem 2011, 118:165–244.
2011, 286:4248–4256.
83. Carter CJ: The fox and the rabbits-environmental variables and population
61. Rival T, Page RM, Chandraratna DS, Sendall TJ, Ryder E, Liu B, Lewis H,
genetics (1) replication problems in association studies and the untapped
Rosahl T, Hider R, Camargo LM, et al: Fenton chemistry and oxidative
power of GWAS (2) vitamin A deficiency, herpes simplex reactivation and
stress mediate the toxicity of the beta-amyloid peptide in a Drosophila
other causes of Alzheimer’s disease. ISRN Neurol 2011, 2011:394678.
model of Alzheimer’s disease. Eur J Neurosci 2009, 29:1335–1347.
84. Rajendran L, Schneider A, Schlechtingen G, Weidlich S, Ries J, Braxmeier T,
62. Wu Z, Du Y, Xue H, Wu Y, Zhou B: Aluminum induces neurodegeneration
Schwille P, Schulz JB, Schroeder C, Simons M, et al: Efficient inhibition of
and its toxicity arises from increased iron accumulation and reactive
the Alzheimer’s disease beta-secretase by membrane targeting.
oxygen species (ROS) production. Neurobiol Aging 2012, 33:199. e191-112.
Science 2008, 320:520–523.
63. Feany MB, Dickson DW: Neurodegenerative disorders with extensive tau
85. Hills ID, Vacca JP: Progress toward a practical BACE-1 inhibitor. Curr Opin
pathology: a comparative study and review. Ann Neurol 1996, 40:139–148.
Drug Discov Devel 2007, 10:383–391.
64. Wittmann CW, Wszolek MF, Shulman JM, Salvaterra PM, Lewis J, Hutton M,
86. Tung JS, Davis DL, Anderson JP, Walker DE, Mamo S, Jewett N, Hom RK,
Feany MB: Tauopathy in Drosophila: neurodegeneration without
Sinha S, Thorsett ED, John V: Design of substrate-based inhibitors of
neurofibrillary tangles. Science 2001, 293:711–714.
human beta-secretase. J Med Chem 2002, 45:259–262.
65. Alonso A, Zaidi T, Novak M, Grundke-Iqbal I, Iqbal K: Hyperphosphorylation
induces self-assembly of tau into tangles of paired helical filaments/
doi:10.1186/1750-1326-8-35
straight filaments. Proc Natl Acad Sci USA 2001, 98:6923–6928.
Cite this article as: Prüßing et al.: Drosophila melanogaster as a model
66. Beharry C, Alaniz ME, Alonso Adel C: Expression of Alzheimer-like organism for Alzheimer’s disease. Molecular Neurodegeneration 2013 8:35.
pathological human Tau induces a behavioral motor and olfactory learning
deficit in Drosophila melanogaster. J Alzheimers Dis 2013, 37:539–550.