Figure 2.
ATFS-1 overexpression induces dopaminergic neurodegenerative phenotypes and cell death in an age-dependent manner. Animals expressing ATFS-1 variant constructs under the Pdat-1 promoter were crossed into animals that express GFP in their dopaminergic neurons (strain BY250; Pdat-1::GFP) as a means to monitor cellular health when ATFS-1 variants are overexpressed. Four neurodegenerative phenotypes were observed: Soma swelling (A–C), dendritic pearling (D–F), neuron cell death (G,H), and dopaminergic signaling reduction (I). A, B, Animals were assayed at days 7 and 10 posthatching for dopaminergic soma swelling phenotypes characterized by enlargement of the cytosolic space surrounding the nucleus of the dopaminergic neuron. Data are shown as the average of three separate transgenic lines with three replicates scored per line per construct. Error bars indicate the SEM of the average of each transgenic line. Statistics were performed using a one-way ANOVA with a Tukey's post hoc test. NS p > 0.05, *p < 0.05. C, Representative images of swollen dopaminergic neuron soma. GFP refers to BY250 (Pdat-1::GFP). Scale bar, 5 μm. D, E, Animals were assayed at days 7 and 10 posthatching for irregular dendritic swelling phenotypes (pearling) characterized by enlargement of the cytosolic space of the dendritic sensillary processes of the CEP dopaminergic neurons. Data are shown as the average of three separate transgenic lines with three replicates per line per construct. Error bars indicate the SEM of the average of each transgenic line. Statistics were performed using a one-way ANOVA with a Tukey's post hoc test. ***p < 0.001. F, Representative images of dendritic pearling in the anterior dopaminergic CEP neurons. GFP-only refers to BY250 (Pdat-1::GFP). Arrows represent irregular GFP puncta. Scale bar, 10 μm. G, H, Animals were assayed at days 7 and 10 posthatching for dopaminergic cell death, characterized by the absence of any of six anterior dopaminergic neurons in part or in whole. Data are shown as the average of three separate transgenic lines with three replicates per line per construct. Error bars indicate the SEM of the average of each transgenic line. Statistics were performed using a one-way ANOVA with a Tukey's post hoc test. *p < 0.05, ***p < 0.001. I, Animals were assayed at day 7 posthatching for basal slowing response activity by measurement of the peristaltic speed on food and off food, expressed as a ratio, and normalized to the ratio quantified for N2 control worms. Data are shown as the average of two transgenic lines using n = 10 animals per each of four replicates per construct. Error bars indicate the SEM of the average. Statistics were performed using a one-way ANOVA with a Tukey's post hoc test. NS, p > 0.05, ***p < 0.001.