Invited reviewModelling motor neuron disease in fruit flies: Lessons from spinal muscular atrophy
Introduction
Motor neuron disease (MND) is an umbrella term for a collection of clinically and aetiologically heterogeneous group of neurological conditions that are characterised by muscle weakness thought to arise from the selective degeneration of lower and/or upper motor neurons. MND causes rapidly progressive motor dysfunction ultimately leading to premature death following respiratory failure in the majority of cases. Genetic factors undoubtedly are major players in disease pathogenesis and progression (Dion et al., 2009, Taylor et al., 2016). This is highlighted by spinal muscular atrophy (SMA), the most common MND striking infants. SMA is typically an autosomal recessive disorder caused by inactivating mutations of the survival motor neuron 1 (SMN1) gene that are partially compensated by the paralogous SMN2 gene. SMA is therefore caused by a reduction rather than total loss of the SMN1/2-encoded SMN protein, with SMN2 copy number influencing age of onset, disease severity and progression rate throughout the lifespan. To this end, SMA is clinically classified as type I (severe), II (intermediate), III (mild) or IV (adult-onset) (Burghes and Beattie, 2009, Kolb and Kissel, 2011). The successful development of a molecular-based therapy for SMA, essentially repairing the SMN2 gene (Talbot and Tizzano, 2017), provides a roadmap for the engineering of therapeutics tailored for other hereditary motor neuron degenerative conditions including the most prevalent MND of adulthood, amyotrophic lateral sclerosis (ALS). Intense research on the molecular genetics underpinning such disorders followed by functional characterisation of the disease-associated genes in animal models are key steps that provide a very good starting point for this challenging endeavour.
The fruit fly Drosophila melanogaster is often the perfect choice of model organism to model neurodegenerative conditions for several reasons (Cauchi and van den Heuvel, 2006, Lessing and Bonini, 2009, McGurk et al., 2015). First, Drosophila has a rapid life cycle with adults emerging after ∼10 days. In addition, its short lifespan (2–3 months) is considered an asset in the context of neurodegenerative disease research. Cost-intensive handling and maintenance, characteristics of vertebrate model organisms do not factor in Drosophila husbandry and ethical restrictions do not apply. Second, since flies were applied to biological research more than a century ago by Thomas Hunt Morgan to discover the role of chromosomes in heredity, a rich genetic toolbox has been developed, allowing researchers to easily manipulate and control Drosophila genetics. A colossal number of mutant and transgenic fly lines have been generated over the years. In view that Drosophila stocks cannot be frozen, they are maintained alive in laboratories all over the world and in ‘public libraries’ including the Bloomington Drosophila Stock Centre (BDSC) at Indiana University (Cook et al., 2010), the Kyoto Stock Centre at the Kyoto Institute of Technology and the Vienna Drosophila Resource Center (Dietzl et al., 2007). Stocks are commonly shared between laboratories or purchased at a low-cost from libraries. Researchers also have access to volumes of curated data on every single Drosophila gene on the FlyBase database (flybase.org). Third, comparison of the fly and human genomes exposed a significant overlap or conservation in genes and pathways (Rubin et al., 2000), with nearly 75% of known human disease-linked genes having homologues in flies (Reiter et al., 2001). In this setting, expression of the disease form of the human gene or mutations in its Drosophila homologue allow researchers to generate fly models of human disease with the aim of discovering mechanisms underpinning the disease process. Furthermore, the compact nature of the Drosophila genome and the relatively lower level of genetic redundancy facilitate exploration of gene function. Fourth, flies are able to respond rapidly to stimuli in addition to performing complex motor behaviours including climbing and flight because they are endowed with a sophisticated neuromuscular system, that though simpler relative to humans, has the basic sensory-motor circuitry, glia and multinucleate muscle fibres (Lloyd and Taylor, 2010). This feature is key for neuromuscular disease modelling and considering the remarkable conservation at a genetic, molecular, and physiological level the fly is increasingly considered as instrumental for gaining insights into the function of novel genes linked to MND in the era of whole-genome sequencing. In this review, we will focus on how Drosophila was harnessed to better understand SMA considering that the strategies undertaken (summarised in Fig. 1) can serve as a paradigm for the unravelling of molecular mechanisms underpinning emerging hereditary neuromuscular conditions.
Section snippets
Loss-of-function approach
Loss-of-function (LOF) gene manipulation is usually the first step undertaken following identification of the Drosophila orthologue of the gene involved in human disease. In Drosophila, genes can be disrupted via various methods including unbiased ethyl methanesulfonate (EMS) mutagenesis, transposable element insertion, insertional/replacement homologous recombination and gene editing via programmable nucleases (reviewed in Ref. Lin et al., 2014). Phenotypes associated with either complete loss
Interaction profiling
Discovery of protein-protein interaction networks is key for unravelling the function of a disease-associated protein and for exploring disease mechanisms. Considering SMN, molecular and structural studies have shown that it operates as part of a large multimeric complex whose components also include a set of diverse proteins, namely Gemins 2–8 and Unrip (Cauchi, 2010). In a recent report, we found that the Drosophila SMN complex is fully conserved, hence, it has the same number of components
Ectopic gene expression
Through the use of the powerful GAL4/UAS system, Drosophila permits investigators to test if expression of human cDNA can correct the phenotypes associated with loss of a homologous fly gene. This was, for instance, the outcome achieved on expression of the ALS-associated TDP-43 gene in mutants lacking the fly equivalent gene (Feiguin et al., 2009), hence indicating functional conservation. The same experiment was not successful for SMN (Borg and Cauchi, 2013), possibly because human SMN has a
Drug discovery
Although Drosophila has traditionally been a ‘powerhouse’ for genetic studies, this premier model organism is finding itself to be central for searching therapeutic compounds that have an ameliorative effect on various diseases including the neurodegenerative spectrum. Compounds that are originally known for their activity in human cells were shown to have the same molecular mechanism of action in Drosophila (Fernandez-Hernandez et al., 2016), a finding that bodes well for the application of
Conclusion & prospects
By focusing on the sterling contributions of the fly model system to the SMA field, in this review we have given a flavour of the types of genetic, molecular and pharmacologic studies that are possible in Drosophila. SMA fly models mirror with remarkable similarities their mammalian counterparts and, importantly, the human condition. Here, we also touched upon several transferable robust assays that were used for SMA modelling (Fig. 2). The power of the fly is unique in that it allows
Acknowledgements
Work in the authors’ laboratory is supported by the University of Malta Research Fund, the Malta Council for Science & Technology Internationalisation Partnership Award and the ALS Malta Foundation.
References (101)
- et al.
CRISPR/Cas9 and genome editing in Drosophila
J. Genet. Genomics
(2014) - et al.
Disruption of snRNP biogenesis factors Tgs1 and pICln induces phenotypes that mirror aspects of SMN-Gemins complex perturbation in Drosophila, providing new insights into spinal muscular atrophy
Neurobiol. Dis.
(2016) - et al.
U bodies respond to nutrient stress in Drosophila
Exp. Cell Res.
(2011) - et al.
Gemin8 is a novel component of the survival motor neuron complex and functions in small nuclear ribonucleoprotein assembly
J. Biol. Chem.
(2006) - et al.
Drosophila SMN complex proteins Gemin2, Gemin3, and Gemin5 are components of U bodies
Exp. Cell Res.
(2010) - et al.
WDR79/TCAB1 plays a conserved role in the control of locomotion and ameliorates phenotypic defects in SMA models
Neurobiol. Dis.
(2017) - et al.
A role for the survival of motor neuron protein in mRNP assembly and transport
Curr. Opin. Neurobiol.
(2016) - et al.
Molecular mechanisms and animal models of spinal muscular atrophy
Biochim. Biophys. Acta
(2015) - et al.
Spinal muscular atrophy: the role of SMN in axonal mRNA regulation
Brain Res.
(2012) - et al.
Depletion of TDP-43 affects Drosophila motoneurons terminal synapsis and locomotive behavior
FEBS Lett.
(2009)
A protein complex network of Drosophila melanogaster
Cell
The power of human protective modifiers: PLS3 and CORO1C unravel impaired endocytosis in spinal muscular atrophy and rescue SMA phenotype
Am. J. Hum. Genet.
SMN is required for sensory-motor circuit function in Drosophila
Cell
The spinal muscular atrophy protein SMN affects Drosophila germline nuclear organization through the U body-P body pathway
Dev. Biol.
Strategies for gene disruption in Drosophila
Cell Biosci.
An SMN-dependent U12 splicing event essential for motor circuit function
Cell
Disruption of SMN function by ectopic expression of the human SMN gene in Drosophila
FEBS Lett.
Gemin2 plays an important role in stabilizing the survival of motor neuron complex
J. Biol. Chem.
A Drosophila model of spinal muscular atrophy uncouples snRNP biogenesis functions of survival motor neuron from locomotion and viability defects
Cell Rep.
Rare missense and synonymous variants in UBE1 are associated with X-linked infantile spinal muscular atrophy
Am. J. Hum. Genet.
Neurocalcin delta suppression protects against spinal muscular atrophy in humans and across species by restoring impaired endocytosis
Am. J. Hum. Genet.
Staufen: a common component of mRNA transport in oocytes and neurons
Trends Cell Biol.
Behavioral and electrophysiological outcomes of tissue-specific Smn knockdown in Drosophila melanogaster
Brain Res.
Axonal transport defects are a common phenotype in Drosophila models of ALS
Hum. Mol. Genet.
The gemin associates of survival motor neuron are required for motor function in Drosophila
PLoS One
GEMINs: potential therapeutic targets for spinal muscular atrophy?
Front. Neurosci.
Genetic interactions between the members of the SMN-Gemins complex in Drosophila
PLoS One
Targeted gene expression as a means of altering cell fates and generating dominant phenotypes
Development
Spinal muscular atrophy: why do low levels of survival motor neuron protein make motor neurons sick?
Nat. Rev. Neurosci.
The fly as a model for neurodegenerative diseases: is it worth the jump?
Neurodegener. Dis.
A motor function for the DEAD-box RNA helicase, Gemin3, in Drosophila
PLoS Genet.
SMN and Gemins: ‘we are family’... or are we? Insights into the partnership between Gemins and the spinal muscular atrophy disease protein SMN
Bioessays
Gem formation upon constitutive Gemin3 overexpression in Drosophila
Cell Biol. Int.
Conserved requirement for DEAD-box RNA helicase Gemin3 in Drosophila oogenesis
BMC Res. Notes
Gem depletion: amyotrophic lateral sclerosis and spinal muscular atrophy crossover
CNS Neurosci. Ther.
Neuromuscular defects in a Drosophila survival motor neuron gene mutant
Hum. Mol. Genet.
Modeling spinal muscular atrophy in Drosophila
PLoS One
Dalfampridine extended release: in multiple sclerosis
CNS Drugs
Deletion of murine SMN exon 7 directed to skeletal muscle leads to severe muscular dystrophy
J. Cell Biol.
New research resources at the Bloomington Drosophila Stock Center
Fly (Austin)
The multiple lives of DEAD-box RNA helicase DP103/DDX20/Gemin3
Biochem. Soc. Trans.
A genetic screen identifies Tor as an interactor of VAPB in a Drosophila model of amyotrophic lateral sclerosis
Biol. Open
A genome-wide transgenic RNAi library for conditional gene inactivation in Drosophila
Nature
Conserved genes act as modifiers of invertebrate SMN loss of function defects
PLoS Genet.
Decreased function of survival motor neuron protein impairs endocytic pathways
Proc. Natl. Acad. Sci. U. S. A.
Genetics of motor neuron disorders: new insights into pathogenic mechanisms
Nat. Rev. Genet.
GAL4 system in Drosophila: a fly geneticist’s Swiss army knife
Genesis
Gemins modulate the expression and activity of the SMN complex
Hum. Mol. Genet.
The translational relevance of Drosophila in drug discovery
EMBO Rep.
Biogenesis of spliceosomal small nuclear ribonucleoproteins
Wiley Interdiscip. Rev. RNA
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