Figure 2.
ER MCSs define the position of OMM and IMM fusion. (A) Representative image of a live U-2 OS cell expressing a marker of the ER (green) and OMM (gray or magenta). Magnified merged time-lapse images show an OMM fusion event (at arrow, middle and right panel) relative to a crossing ER tubule (at arrow, in bottom panels). (B) Linescan analysis (dashed line) at t = 20 s from A plots mean FI of an ER tubule that crosses over the location of fusion. (C) Percentage of OMM fusion events occurring at ER tubules relative to the mean coverage of ER along mitochondria (n = 20 events in seven cells). (D) As in A for cells expressing a marker of the ER (green) and mitochondrial matrix (magenta). Magnified merged time-lapse images show an IMM fusion event (at arrow, middle and bottom panel) relative to a crossing ER tubule (green, bottom panels). (E) As in B for 50-s panel in D. (F) As in C for D (n = 20 events in 17 cells). (G) Representative image of cells expressing an ER (green) and photoconvertible OMM marker (gray). Photoconversion of individual mitochondria converts gray mitochondria to magenta. Conversely, bona fide OMM fusion between magenta and gray mitochondria leads to content/color mixing (compare 84 s with 96 s). (H) Cartoon demonstrating content mixing of OMM upon fusion. (I) Percentage of OMM fusion events scored by content mixing that occur at a crossing ER tubule relative to the average percentage coverage of ER along mitochondria (n = 30 events in 14 cells). (J) As in G for a photoconvertible matrix marker. Time lapse images showing IMM fusion confirmed by content mixing (between 9 s and 21 s). (K) Cartoon demonstrating content mixing of matrix upon fusion. (L) As in I for matrix content mixing experiment in J (n = 33 events in 10 cells). Scale bars for whole cell = 5 µm; insets, 1 µm.

ER MCSs define the position of OMM and IMM fusion. (A) Representative image of a live U-2 OS cell expressing a marker of the ER (green) and OMM (gray or magenta). Magnified merged time-lapse images show an OMM fusion event (at arrow, middle and right panel) relative to a crossing ER tubule (at arrow, in bottom panels). (B) Linescan analysis (dashed line) at t = 20 s from A plots mean FI of an ER tubule that crosses over the location of fusion. (C) Percentage of OMM fusion events occurring at ER tubules relative to the mean coverage of ER along mitochondria (n = 20 events in seven cells). (D) As in A for cells expressing a marker of the ER (green) and mitochondrial matrix (magenta). Magnified merged time-lapse images show an IMM fusion event (at arrow, middle and bottom panel) relative to a crossing ER tubule (green, bottom panels). (E) As in B for 50-s panel in D. (F) As in C for D (n = 20 events in 17 cells). (G) Representative image of cells expressing an ER (green) and photoconvertible OMM marker (gray). Photoconversion of individual mitochondria converts gray mitochondria to magenta. Conversely, bona fide OMM fusion between magenta and gray mitochondria leads to content/color mixing (compare 84 s with 96 s). (H) Cartoon demonstrating content mixing of OMM upon fusion. (I) Percentage of OMM fusion events scored by content mixing that occur at a crossing ER tubule relative to the average percentage coverage of ER along mitochondria (n = 30 events in 14 cells). (J) As in G for a photoconvertible matrix marker. Time lapse images showing IMM fusion confirmed by content mixing (between 9 s and 21 s). (K) Cartoon demonstrating content mixing of matrix upon fusion. (L) As in I for matrix content mixing experiment in J (n = 33 events in 10 cells). Scale bars for whole cell = 5 µm; insets, 1 µm.

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