Figure 4.
Endosomes remain close to MTs. (a) Spinning disk + SRRF image of MTs (green) and dextran vesicles (magenta) in live cells. The white arrowheads indicate representative vesicles on the MT. (b) Spinning disk + SRRF image of p62 (top left), Rab5 vesicles (top right) in live cells, and their merge (bottom, p62 in green and Rab5 in magenta). The white arrowheads in the merged image indicate representative vesicles that colocalize with p62 and are shown as green and magenta arrowheads in the p62 and Rab5 images, respectively. In a and b, “N” marks the location/direction of the nucleus. (c) Plot of the probability of co-occurrence of p62 with Rab5. n = ∼40 cells across two independent repeats. (d) Fluorescence images of EGF (left), p62 (middle), and p150 merge (right) obtained using Airyscan confocal microscopy. (e) The insets marked with a white box in d are depicted; EGF, p62, and p150 channel insets are depicted as intensity maps and the white arrowheads point to EGF punctae, some of which colocalize with p62 and p150. (f) Plot of the probability of co-occurrence of EGF with p62 and p150, indicating a high likelihood of dynactin being present in a complex with endosomal cargo. n = ∼60 cells across three independent repeats. (g) Overlay of confocal images of MT (green) and dextran vesicles (magenta), and EM images of the same cell (gray). (h) EM image of the region indicated with the white square in g. (i) Merge of h and confocal fluorescence image of dextran (magenta) of the region in h showing a representative MT (dashed white line) and dextran endosome (dashed white circle). (j) Overlay of confocal images of MT (green) and EGF vesicles (magenta), and EM images of the same cell (gray). (k) EM image of the region indicated with the white square in j. (l) Merge of k and confocal fluorescence image of EGF (magenta) of the region in k showing a representative MT (dashed white line) and EGF endosome (dashed white circle). (m) Quantification of the measured distance between dextran (“Dex”) and EGF endosomes and MTs. “n.s.” indicates no significant difference (P = 0.3), one-way ANOVA, Tukey Kramer post-hoc test. n = 6 cells from two independent repeats. Error bars represent SD.

Endosomes remain close to MTs. (a) Spinning disk + SRRF image of MTs (green) and dextran vesicles (magenta) in live cells. The white arrowheads indicate representative vesicles on the MT. (b) Spinning disk + SRRF image of p62 (top left), Rab5 vesicles (top right) in live cells, and their merge (bottom, p62 in green and Rab5 in magenta). The white arrowheads in the merged image indicate representative vesicles that colocalize with p62 and are shown as green and magenta arrowheads in the p62 and Rab5 images, respectively. In a and b, “N” marks the location/direction of the nucleus. (c) Plot of the probability of co-occurrence of p62 with Rab5. n = ∼40 cells across two independent repeats. (d) Fluorescence images of EGF (left), p62 (middle), and p150 merge (right) obtained using Airyscan confocal microscopy. (e) The insets marked with a white box in d are depicted; EGF, p62, and p150 channel insets are depicted as intensity maps and the white arrowheads point to EGF punctae, some of which colocalize with p62 and p150. (f) Plot of the probability of co-occurrence of EGF with p62 and p150, indicating a high likelihood of dynactin being present in a complex with endosomal cargo. n = ∼60 cells across three independent repeats. (g) Overlay of confocal images of MT (green) and dextran vesicles (magenta), and EM images of the same cell (gray). (h) EM image of the region indicated with the white square in g. (i) Merge of h and confocal fluorescence image of dextran (magenta) of the region in h showing a representative MT (dashed white line) and dextran endosome (dashed white circle). (j) Overlay of confocal images of MT (green) and EGF vesicles (magenta), and EM images of the same cell (gray). (k) EM image of the region indicated with the white square in j. (l) Merge of k and confocal fluorescence image of EGF (magenta) of the region in k showing a representative MT (dashed white line) and EGF endosome (dashed white circle). (m) Quantification of the measured distance between dextran (“Dex”) and EGF endosomes and MTs. “n.s.” indicates no significant difference (P = 0.3), one-way ANOVA, Tukey Kramer post-hoc test. n = 6 cells from two independent repeats. Error bars represent SD.

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