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Dominant, toxic gain-of-function mutations in gars lead to non-cell autonomous neuropathology.

Grice SJ, Sleigh JN, Motley WW, Liu JL, Burgess RW, Talbot K, Cader MZ - Hum. Mol. Genet. (2015)

Bottom Line: Intriguingly, neuronal toxicity is, at least in part, non-cell autonomous, as expression of mutant GlyRS in mesoderm or muscle alone results in similar pathology.This mutant GlyRS toxic gain-of-function, which is WHEP domain-dependent, coincides with abnormal NMJ assembly, leading to synaptic degeneration, and, ultimately, reduced viability.Our findings suggest that mutant GlyRS gains access to ectopic sub-compartments of the motor neuron, providing a possible explanation for the selective neuropathology caused by mutations in a widely expressed gene.

View Article: PubMed Central - PubMed

Affiliation: MRC Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, South Parks Road, Oxford OX1 3PT, UK.

No MeSH data available.


Related in: MedlinePlus

garsP234KY expression causes NMJ defects. NMJs consist of boutons that grow in lines resembling beads on a string (arrowheads, A). Nerves were visualized with anti-HRP (red) and the post-synaptic apparatus with anti-DLG (green). (A) Representative NMJs from muscles 6 and 7 of control (1032-GAL4) and ubiquitous garsP234KY (1032-GAL4; UAS-garsP234KY) larvae at the late L3 stage. (B) Ubiquitous, mesodermal (how-GAL4) and muscle (MHC-GAL4) expression of garsP234KY caused a reduction in bouton number. (C) Ubiquitous, mesodermal and muscle expression also lead to an increase in bouton size. Neuronal garsP234KY expression (elav-GAL4) caused no overt synaptic phenotype. For all graphs, *P < 0.05, **P < 0.01, ***P < 0.001 Bonferroni's/Dunn's multiple comparison test. At least 20 larvae were scored per NMJ. Scale bars = 10 µm.
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DDV176F2: garsP234KY expression causes NMJ defects. NMJs consist of boutons that grow in lines resembling beads on a string (arrowheads, A). Nerves were visualized with anti-HRP (red) and the post-synaptic apparatus with anti-DLG (green). (A) Representative NMJs from muscles 6 and 7 of control (1032-GAL4) and ubiquitous garsP234KY (1032-GAL4; UAS-garsP234KY) larvae at the late L3 stage. (B) Ubiquitous, mesodermal (how-GAL4) and muscle (MHC-GAL4) expression of garsP234KY caused a reduction in bouton number. (C) Ubiquitous, mesodermal and muscle expression also lead to an increase in bouton size. Neuronal garsP234KY expression (elav-GAL4) caused no overt synaptic phenotype. For all graphs, *P < 0.05, **P < 0.01, ***P < 0.001 Bonferroni's/Dunn's multiple comparison test. At least 20 larvae were scored per NMJ. Scale bars = 10 µm.

Mentions: As motor defects were present at larval stages in mutant flies, we looked for changes in the structure and innervation pattern of the larval NMJ. All subsequent experiments were performed at 25°C, as significant toxicity is observed at this temperature in all the tissues studied. Abdominal ventrolateral muscles 6 and 7 are innervated in a stereotyped manner in Drosophila larvae by a defasciculated segmental nerve d. Wild-type NMJs grow by adding circular synaptic specializations called boutons, which form structures like beads on a string, where both pre- and post-synaptic regions closely overlap (Fig. 2A). Larvae ubiquitously expressing garsP234KY displayed reduced bouton number and increased bouton size when compared with wild-type and garsWT-overexpressing flies (Fig. 2B and C). No gross changes in muscle size or structure were observed in any of the genotypes analysed (Supplementary Material, Fig. S2). Again, tissue-specific GlyRSP234KY toxicity was analysed by expressing garsP234KY in mesoderm (how-GAL4), muscle (MHC-GAL4) or neurons (elav-GAL4), whereas flies ubiquitously expressing garsWT were used as controls. Bouton number defects were also observed in larvae with mesoderm and muscle-driven GlyRSP234KY, whereas neuronal garsP234KY expression caused no overt synaptic phenotype (Fig. 2B and C).Figure 2.


Dominant, toxic gain-of-function mutations in gars lead to non-cell autonomous neuropathology.

Grice SJ, Sleigh JN, Motley WW, Liu JL, Burgess RW, Talbot K, Cader MZ - Hum. Mol. Genet. (2015)

garsP234KY expression causes NMJ defects. NMJs consist of boutons that grow in lines resembling beads on a string (arrowheads, A). Nerves were visualized with anti-HRP (red) and the post-synaptic apparatus with anti-DLG (green). (A) Representative NMJs from muscles 6 and 7 of control (1032-GAL4) and ubiquitous garsP234KY (1032-GAL4; UAS-garsP234KY) larvae at the late L3 stage. (B) Ubiquitous, mesodermal (how-GAL4) and muscle (MHC-GAL4) expression of garsP234KY caused a reduction in bouton number. (C) Ubiquitous, mesodermal and muscle expression also lead to an increase in bouton size. Neuronal garsP234KY expression (elav-GAL4) caused no overt synaptic phenotype. For all graphs, *P < 0.05, **P < 0.01, ***P < 0.001 Bonferroni's/Dunn's multiple comparison test. At least 20 larvae were scored per NMJ. Scale bars = 10 µm.
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Related In: Results  -  Collection

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DDV176F2: garsP234KY expression causes NMJ defects. NMJs consist of boutons that grow in lines resembling beads on a string (arrowheads, A). Nerves were visualized with anti-HRP (red) and the post-synaptic apparatus with anti-DLG (green). (A) Representative NMJs from muscles 6 and 7 of control (1032-GAL4) and ubiquitous garsP234KY (1032-GAL4; UAS-garsP234KY) larvae at the late L3 stage. (B) Ubiquitous, mesodermal (how-GAL4) and muscle (MHC-GAL4) expression of garsP234KY caused a reduction in bouton number. (C) Ubiquitous, mesodermal and muscle expression also lead to an increase in bouton size. Neuronal garsP234KY expression (elav-GAL4) caused no overt synaptic phenotype. For all graphs, *P < 0.05, **P < 0.01, ***P < 0.001 Bonferroni's/Dunn's multiple comparison test. At least 20 larvae were scored per NMJ. Scale bars = 10 µm.
Mentions: As motor defects were present at larval stages in mutant flies, we looked for changes in the structure and innervation pattern of the larval NMJ. All subsequent experiments were performed at 25°C, as significant toxicity is observed at this temperature in all the tissues studied. Abdominal ventrolateral muscles 6 and 7 are innervated in a stereotyped manner in Drosophila larvae by a defasciculated segmental nerve d. Wild-type NMJs grow by adding circular synaptic specializations called boutons, which form structures like beads on a string, where both pre- and post-synaptic regions closely overlap (Fig. 2A). Larvae ubiquitously expressing garsP234KY displayed reduced bouton number and increased bouton size when compared with wild-type and garsWT-overexpressing flies (Fig. 2B and C). No gross changes in muscle size or structure were observed in any of the genotypes analysed (Supplementary Material, Fig. S2). Again, tissue-specific GlyRSP234KY toxicity was analysed by expressing garsP234KY in mesoderm (how-GAL4), muscle (MHC-GAL4) or neurons (elav-GAL4), whereas flies ubiquitously expressing garsWT were used as controls. Bouton number defects were also observed in larvae with mesoderm and muscle-driven GlyRSP234KY, whereas neuronal garsP234KY expression caused no overt synaptic phenotype (Fig. 2B and C).Figure 2.

Bottom Line: Intriguingly, neuronal toxicity is, at least in part, non-cell autonomous, as expression of mutant GlyRS in mesoderm or muscle alone results in similar pathology.This mutant GlyRS toxic gain-of-function, which is WHEP domain-dependent, coincides with abnormal NMJ assembly, leading to synaptic degeneration, and, ultimately, reduced viability.Our findings suggest that mutant GlyRS gains access to ectopic sub-compartments of the motor neuron, providing a possible explanation for the selective neuropathology caused by mutations in a widely expressed gene.

View Article: PubMed Central - PubMed

Affiliation: MRC Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, South Parks Road, Oxford OX1 3PT, UK.

No MeSH data available.


Related in: MedlinePlus