Figure 5. Capture of small vesicles by... | Rockefeller University Press

$Figure 5. Capture of small vesicles by rerouting TPD54 to mitochondria. (A) Fluorescence microscopy images of mCherry-FKBP-TPD54 in HeLa cells. Cells expressing mCherry-FKBP-TPD54 and dark MitoTrap were fixed after no rapamycin application (Ctrl) or after 20 s, 5 min, or 30 min of rapamycin addition (200 nM). The pictured cell was then imaged by EM. Scale bar, 10 µm. (B) Sample electron micrographs of the cells shown in A. Insets, 3× zoom. Scale bars, 200 nm, 50 nm (insets). (C) Segmented view of mitochondria (gray) and vesicles (purple) in the images shown in B. (D) Profiles of segmented vesicles from electron micrographs. All vesicles segmented from the control dataset are shown with a random sample from the treatment groups as a comparison. The sample size is in proportion to the capture of vesicles at the mitochondria (34, 320, 594, and 1,347 for control, 20 s, 5 min, and 30 min, respectively). (E) Left: violin plot to show the diameter of vesicles imaged in each dataset. Spots represent individual vesicles. Marker shows the median. Center: box plot to show the number of vesicles captured per 1 µm of mitochondrial membrane. Spots show the number per micrograph in each dataset. Boxes show the interquartile range and median, and whiskers show 9th and 91st percentiles. Right: violin plot to show the fraction of mitochondrial membrane that is decorated with vesicles. Spots show individual mitochondria from the dataset. Time, minutes and seconds. See Materials and methods for details.$

Figure 5.

Capture of small vesicles by rerouting TPD54 to mitochondria. (A) Fluorescence microscopy images of mCherry-FKBP-TPD54 in HeLa cells. Cells expressing mCherry-FKBP-TPD54 and dark MitoTrap were fixed after no rapamycin application (Ctrl) or after 20 s, 5 min, or 30 min of rapamycin addition (200 nM). The pictured cell was then imaged by EM. Scale bar, 10 µm. (B) Sample electron micrographs of the cells shown in A. Insets, 3× zoom. Scale bars, 200 nm, 50 nm (insets). (C) Segmented view of mitochondria (gray) and vesicles (purple) in the images shown in B. (D) Profiles of segmented vesicles from electron micrographs. All vesicles segmented from the control dataset are shown with a random sample from the treatment groups as a comparison. The sample size is in proportion to the capture of vesicles at the mitochondria (34, 320, 594, and 1,347 for control, 20 s, 5 min, and 30 min, respectively). (E) Left: violin plot to show the diameter of vesicles imaged in each dataset. Spots represent individual vesicles. Marker shows the median. Center: box plot to show the number of vesicles captured per 1 µm of mitochondrial membrane. Spots show the number per micrograph in each dataset. Boxes show the interquartile range and median, and whiskers show 9th and 91st percentiles. Right: violin plot to show the fraction of mitochondrial membrane that is decorated with vesicles. Spots show individual mitochondria from the dataset. Time, minutes and seconds. See Materials and methods for details.

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