Motor neurons and glia exhibit specific individualized responses to TDP-43 expression in a Drosophila model of amyotrophic lateral sclerosis

Abstract

SUMMARYAmyotrophic lateral sclerosis (ALS) is a fatal disease characterized by complex neuronal and glial phenotypes. Recently, RNA-based mechanisms have been linked to ALS via RNA-binding proteins such as TDP-43, which has been studied in vivo using models ranging from yeast to rodents. We have developed a Drosophila model of ALS based on TDP-43 that recapitulates several aspects of pathology, including motor neuron loss, locomotor dysfunction and reduced survival. Here we report the phenotypic consequences of expressing wild-type and four different ALS-linked TDP-43 mutations in neurons and glia. We show that TDP-43-driven neurodegeneration phenotypes are dose- and age-dependent. In motor neurons, TDP-43 appears restricted to nuclei, which are significantly misshapen due to mutant but not wild-type protein expression. In glia and in the developing neuroepithelium, TDP-43 associates with cytoplasmic puncta. TDP-43-containing RNA granules are motile in cultured motor neurons, although wild-type and mutant variants exhibit different kinetic properties. At the neuromuscular junction, the expression of TDP-43 in motor neurons versus glia leads to seemingly opposite synaptic phenotypes that, surprisingly, translate into comparable locomotor defects. Finally, we explore sleep as a behavioral readout of TDP-43 expression and find evidence of sleep fragmentation consistent with hyperexcitability, a suggested mechanism in ALS. These findings support the notion that although motor neurons and glia are both involved in ALS pathology, at the cellular level they can exhibit different responses to TDP-43. In addition, our data suggest that individual TDP-43 alleles utilize distinct molecular mechanisms, which will be important for developing therapeutic strategies.

mRNP granules

“…This is particularly true of most P-body and stress granule factors 10 (with exceptions e.g., Dcp2 in P-bodies). Germ granule components (e.g., Vasa in Drosophila nuage; residency time of 60s; 126 PGL-1 in C. elegans perinuclear P-granules 20s 21 ), and neuronal granule components (e.g., Staufen in Drosophila neuronal granules; residency > 10 min; 104 TDP-43 in Drosophila neuronal granules 10s 127 ) also exhibit a wide range of shuttling rates. More telling are FRAP studies on fluorescently labeled mRNAs, which have indicated that both rapidly exchanging and immobile fractions of a single mRNA species can exist within stress granules simultaneously.…”

“…147 In addition, pathogenic forms of TDP-43 expressed in C. elegans or Drosophila show slower exchange rates from aggregates or neuronal granules by FRAP. 127,148 Several reasons have been proposed as to why persistent stress granules or related mRNP aggregates could be toxic to cells. 136,144 Sequestration of protein factors in granules could lead to a reduced ability of granules to appropriately remodel mRNPs, and may limit the available pool of a protein that also functions elsewhere in the cell.…”

Glial cells in amyotrophic lateral sclerosis

“…The authors suggested that changes in the Drosophila orthologues for EAAT1/EAAT2 which are involved in glutamate transport, might underlie the phenotype in both knockdown as well as overexpression models. Although motor neuron specific mTDP-43 expression in Drosophila seems to confer different effects on the cellular level as compared to mTDP-43 expression in glial cells, its effects on the phenotypical level is very similar (Estes et al, 2013). …”

A fruitful endeavor: Modeling ALS in the fruit fly

“…The results have shown that WT hTDP-43, as well as A315T, D169G, G298S and N345K mutants, mislocalize to the cytoplasm when overexpressed (Estes et al, 2013). WT and mutant hTDP-43 overexpression in both glial and motor neuron cells also reduces bouton number at larval NMJ’s and hinders larval motor function, specifically their ability to turn over after being flipped upside down.…”