Figure S2.
Other domain components are transported independently in the anterograde direction and substantially cotransported in the retrograde direction. (A) Representative micrographs captured from live images show transport of Navβ2-mKate2 versus Navβ1-EGFP (top row) and Navβ1-mKate2 versus KCNQ3-EGFP (bottom row) in the anterograde direction. White arrows indicate examples of vesicles that are only mKate2 labeled. Insets show higher power images of vesicles indicated by the arrows. Time of imaging (seconds) is indicated in each panel. (B) Corresponding duplicate kymographs of anterogradely transported vesicles, indicated by the white arrows in A, are shown circled and their trajectories are indicated by the dotted blue arrows. Scale bars, 1 µm. (C) Quantification of cotransport for the constructs indicated. In the anterograde direction (top panel), no cotransport was observed for Navβ2-mKate2 with Navβ1-EGFP (15 axons, 97 vesicles) or KCNQ3-mKate2 with Navβ1-EGFP (15 axons, 96 vesicles). There was substantial cotransport in the retrograde direction (bottom panel) for Navβ2-EGFP with Navβ1-mKate2 (56.8% ± 4.7%, n = 15 axons, 144 vesicles) and KCNQ3-EGFP with Navβ1-mKate2 (45.8% ± 5.0%, n = 14 axons, 122 vesicles). Error bars indicate mean ± SEM. (D) Quantification of cotransport for the constructs indicated. In the anterograde direction (top panel), no cotransport was observed for Navβ1-mKate2 with KCNQ3-EGFP (14 axons, 96 vesicles), for NrCAM-mKate2 with KCNQ3-EGFP (14 axons, 76 vesicles), or for Caspr-EGFP with KCNQ3-mKate2 (21 axons, 96 vesicles). In the retrograde direction (bottom panel), there was substantial cotransport for Navβ2-mKate2 with Navβ1-EGFP (56.8% ± 4.7%, n = 15 axons, 144 vesicles), for KCNQ3-EGFP with Navβ1-mKate2 (45.8% ± 5.0%, n = 14 axons, 122 vesicles), and for Caspr-EGFP with KCNQ3-mKate2 (48.6% ± 5.1%, n = 21 axons, 166 vesicles). Error bars indicate mean ± SEM. (E) Quantification of cotransport for the constructs indicated. In the anterograde direction (top panel), no cotransport was observed for NrCAM-mKate2 with Caspr-EGFP (15 axons, 72 vesicles) or for KCNQ3-mKate2 with Caspr-EGFP (21 axons, 96 vesicles). In the retrograde direction (bottom panel), there was substantial cotransport for NrCAM-mKate2 with Caspr-EGFP (49.2% ± 4.2%, n = 15 axons, 118 vesicles) and for KCNQ3-mKate2 with Caspr-EGFP (54.8% ± 5.7%, n = 21 axons, 166 vesicles). Error bars indicate mean ± SEM. (F) An example of NF186-EGFP–containing vesicle that colabeled with Lysotracker is indicated by the yellow arrow in the micrograph (left) and circled in the kymograph (right); its retrograde trajectory is indicated by the dotted red arrow. Axon numbers analyzed: n = 10.

Other domain components are transported independently in the anterograde direction and substantially cotransported in the retrograde direction. (A) Representative micrographs captured from live images show transport of Navβ2-mKate2 versus Navβ1-EGFP (top row) and Navβ1-mKate2 versus KCNQ3-EGFP (bottom row) in the anterograde direction. White arrows indicate examples of vesicles that are only mKate2 labeled. Insets show higher power images of vesicles indicated by the arrows. Time of imaging (seconds) is indicated in each panel. (B) Corresponding duplicate kymographs of anterogradely transported vesicles, indicated by the white arrows in A, are shown circled and their trajectories are indicated by the dotted blue arrows. Scale bars, 1 µm. (C) Quantification of cotransport for the constructs indicated. In the anterograde direction (top panel), no cotransport was observed for Navβ2-mKate2 with Navβ1-EGFP (15 axons, 97 vesicles) or KCNQ3-mKate2 with Navβ1-EGFP (15 axons, 96 vesicles). There was substantial cotransport in the retrograde direction (bottom panel) for Navβ2-EGFP with Navβ1-mKate2 (56.8% ± 4.7%, n = 15 axons, 144 vesicles) and KCNQ3-EGFP with Navβ1-mKate2 (45.8% ± 5.0%, n = 14 axons, 122 vesicles). Error bars indicate mean ± SEM. (D) Quantification of cotransport for the constructs indicated. In the anterograde direction (top panel), no cotransport was observed for Navβ1-mKate2 with KCNQ3-EGFP (14 axons, 96 vesicles), for NrCAM-mKate2 with KCNQ3-EGFP (14 axons, 76 vesicles), or for Caspr-EGFP with KCNQ3-mKate2 (21 axons, 96 vesicles). In the retrograde direction (bottom panel), there was substantial cotransport for Navβ2-mKate2 with Navβ1-EGFP (56.8% ± 4.7%, n = 15 axons, 144 vesicles), for KCNQ3-EGFP with Navβ1-mKate2 (45.8% ± 5.0%, n = 14 axons, 122 vesicles), and for Caspr-EGFP with KCNQ3-mKate2 (48.6% ± 5.1%, n = 21 axons, 166 vesicles). Error bars indicate mean ± SEM. (E) Quantification of cotransport for the constructs indicated. In the anterograde direction (top panel), no cotransport was observed for NrCAM-mKate2 with Caspr-EGFP (15 axons, 72 vesicles) or for KCNQ3-mKate2 with Caspr-EGFP (21 axons, 96 vesicles). In the retrograde direction (bottom panel), there was substantial cotransport for NrCAM-mKate2 with Caspr-EGFP (49.2% ± 4.2%, n = 15 axons, 118 vesicles) and for KCNQ3-mKate2 with Caspr-EGFP (54.8% ± 5.7%, n = 21 axons, 166 vesicles). Error bars indicate mean ± SEM. (F) An example of NF186-EGFP–containing vesicle that colabeled with Lysotracker is indicated by the yellow arrow in the micrograph (left) and circled in the kymograph (right); its retrograde trajectory is indicated by the dotted red arrow. Axon numbers analyzed: n = 10.

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