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. 2022 Jan 24;11:e74387. doi: 10.7554/eLife.74387

Figure 4. Endocytic accessory proteins are associated with the PSD.

(A) Example images of dendrites expressing Homer1c-mCherry to mark PSDs and GFP-CLCa visualized as black and white images (left, middle panel), and merge (right panel). (B) Example images of neurons co-expressing tagged endocytic proteins relative to Homer1c. (A–B) Scale bars: 5 µm, zoom dimensions: 1 µm. (C) Percentage of synapses associated with endocytic proteins, represented as mean ± SEM. Relative to Homer1c-CLCa association (N = 8), PICALM-mCherry (N = 6, p < 0.05), FCHO1-mCherry (N = 6, p < 0.001), Epsn2-mCherry (N = 9, p < 0.01), and Amph-mCherry (N = 6, p < 0.01) were significantly less often associated with the PSD, while β2-adaptin- GFP (N = 6), GFP-Eps15 (N = 5), GFP-Itsn1L (N = 5), HIP1R-GFP (N = 9), GFP-CPG2 (N = 5), Dyn2-GFP (N = 5) were not different from GFP-CLCa. (D) Example images of neurons expressing Homer1c-ALFA labeled with Cy3 (magenta) and endogenously GFP-tagged endocytic proteins using CRISPR/Cas9-mediated genome editing (green). Scale bar: 5 µm, zoom: 500 nm. Data represented as mean ± SEM.

Figure 4—source data 1. Excel sheet with numerical data represented as plots Figure 4C.

Figure 4.

Figure 4—figure supplement 1. Endogenous labeling of endocytic proteins in neurons.

Figure 4—figure supplement 1.

(A) Quantification of the average number of EZ proteins puncta per synapse after overexpression (OE) or knock-in (KI). OE data represented is a copy from the data shown in Figure 4C. KI Eps15 (N = 8), KI Itsn1 (N = 9), KI Dyn2 (N = 7). Data is plotted as mean ± SEM. (B) Example images (confocal) of Homer1c-ALFA labeled with Cy3 and endogenous labeling of EZ proteins using antibodies. Scale bar: 5 µm, zoom: 500 nm.
Figure 4—figure supplement 1—source data 1. Excel sheet with numerical data represented as a plot in Figure 4—figure supplement 1A.