Fig. 3.

Altered localization of presynaptic endocytic proteins in smn-1 loss-of-function animals. (A–E) Representative images of fluorescently tagged, presynaptic proteins expressed in the dorsal nerve cord of cholinergic DA motor neurons of smn-1(+) control, smn-1(ok355), and smn-1(rt248) animals. (Scale bar, 10 μm.) (F) Percent change from control for SYD-2 (α-liprin), NLP-21 (GGARAF neuropeptide family), APT-4 (AP2 α-adaptin), ITSN-1 (DAP160/Intersectin), and SNB-1 (synaptobrevin) in smn-1(ok355) animals is reported for average puncta width, puncta total intensity, and linear density (number per micrometer). ITSN-1 and SNB-1 are reported for smn-1(rt248). Light and dark shading indicate an increase or decrease, respectively, in the smn-1(ok355) and smn-1(rt248) animals compared with smn-1(+) control (see Dataset S1 for extended analysis). Mann–Whitney U test, two-tailed: *P < 0.05; **P < 0.01; ***P < 0.001. (G–I) A single copy of the [smn-1(+)] construct rtSi10 fully rescued APT-4 linear density [control smn-1(+) animals vs. rescued smn-1(ok355);rtSi10[smn-1(+)] P = 0.5; smn-1(ok355) vs. rescued P = 0.002], partially ameliorated puncta width defects [control smn-1(+) vs. rescued P = 0.002; smn-1(ok355) vs. rescued P = 0.03], but may not have improved total intensity [control smn-1(+) vs. rescued P = 5e-04; smn-1(ok355) vs. rescued P = 0.09]. Distributions of APT-4 puncta width, linear density, and puncta total intensity in smn-1(ok355) animals are compared with distributions in control smn-1(+) and rescued animals. Results are presented as kernel density estimates, which convert distribution histograms into smooth, continuous density function curves. The x-axis values were log2-transformed before the calculation of the density function. Linear density (puncta per micrometer) values are less than 1, resulting in negative values after log2 transform (Dataset S1). At least three independent trials were performed (n > 25 animals in total/genotype).