Figure 6.

Airyscan FAST imaging of live galactose adapted mCherry-EGFP-SYNJ2B-TM H9c2 cells stained with LysoView 650 reveals transient contact and transfer between the mitochondrial network and red-only dots. (A) Full frame of video S3 at time 0:35 with zoomed-in area indicated (white box) for the selected time points (small panels, right). White arrows in the full frame show examples of red-only dots in close proximity with tubular mitochondria. White arrowheads in the small panels highlight a red-only dot approaching and contacting the mitochondrial network, producing a brief colocalized signal (yellow arrowheads) around time 2:34. At time 2:55, EGFP fluorescence has diminished, and the structure again appears red-only (white arrowheads). In subsequent frames, small protrusions of EGFP fluorescence (notched yellow arrowheads at time points 4:00 and 4:08) can be seen upon close inspection to extend from the mitochondrion and into the red-only dot, and then rapidly disappear (see also video S3). (B) Full frame of video S4 at time 1:47 with zoomed-in area indicated (white box) as in (A). White arrowheads indicate a red-only dot approaching the mitochondrial network from time point 2:07. The EGFP signal remains detectable within the red-only dot for a few seconds around time 2:19 (yellow arrowheads) and then disappears around time point 2:22 (see also video S4). (C) Full frame of video S5 at time 0:06 with zoomed-in area indicated (white box) as in (A) and (B). White arrowheads indicate a red-only dot which encounters a tubular mitochondrion from time point 0:24 (yellow arrowheads). In subsequent frames, small protrusions of EGFP fluorescence can be seen to extend into the red-only dots (notched yellow arrowheads at time 1:36, 1:55, 2:03, and 2:12). An enlarged partial view (white boxes) of the mCherry and EGFP channels is shown below each time point. Scale bars: 10 μm (full frames), 2 μm (small panels), and 1 μm (enlarged views).