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. Author manuscript; available in PMC: 2024 May 3.
Published in final edited form as: Neuron. 2023 Mar 16;111(9):1381–1390.e6. doi: 10.1016/j.neuron.2023.02.029

Figure 3. Partial loss of TopoII activity suppresses poly(GR) toxicity in flies and neurodegeneration in C9ORF72 iPSC-derived motor neurons.

Figure 3.

(A) Genetic knockdown of TopoII strongly suppresses poly(GR) toxicity in Drosophila photoreceptor neuron. Representative images of adult external and internal eye phenotypes of flies with different genotype. UAS-GFP was used as control for genetic crosses with other UAS elements. Scale bar 25 uM.

(B) Quantification of the effects of TopoII knockdown on poly(GR)-induced external eye degeneration phenotype. The number of flies of each genotype is presented in each column. The P value was determined by chi-square test.

(C) Doxorubicin suppresses poly(GR)-induced toxicity in Drosophila eye. Representative images of eye phenotypes of control and (GR)80 flies fed vehicle or doxorubicin.

(D) Quantification of the eye degeneration phenotype. The number of flies of each genotype is shown in each column. The P value was determined by chi-square test.

(E) Pan-neuronal downregulation of TopoII or Hsp90 rescues the locomotor defect of (GR)80 flies in negative geotaxis climbing assay. For each genotype, climbing assay was performed at least five cohorts consisting of 16-20 flies grown at 27 °C at 3 weeks (n=5, 7, 8, and 8, Tukey–Kramer test). UAS-mCherry RNAi was used as control for genetic crosses with other UAS-RNAi elements.

(F) Quantification of the efficiency of TopoIIβ ASOs knockdown in iPSC-derived motor neurons. One-way ANOVA with Dunnett’s post-hoc test for multiple comparisons.

(G) Decreased survival of C9ORF72 ALS patient motor neurons. Three control and three C9ORF72 iPSC lines were differentiated (n = 100 neurons per line).

(H) Decreased survival of C9ORF72 motor neurons was significantly increased by TopoIIβ ASOs treatment. Three C9ORF72 iPSC lines were differentiated (n = 100 neurons per line).

(I) Survival of control iPSC-derived motor neurons treated with TopoIIβ ASOs or control ASO. Three iPSC lines were differentiated (n = 100 neurons per line). All motor neuron survival experiments were analyzed by two-sided log-rank test, and statistical significance was calculated from the entire survival time course. All data values are mean ± s.e.m. **** P < 0.0001, *** P < 0.001, **P < 0.01, * P < 0.05, ns, not significant.

See also Figure S3.