Figure 4. Dynamics of the vacuolar membrane during the incorporation of Rubisco-containing bodies (RCBs).

Leaves accumulating the chloroplast stroma marker RBCS-mRFP along with the vacuolar membrane marker VHP1-mGFP were incubated in sugar-free solution in darkness for 6–8 hr from dawn, and the behavior of cytosolic RCBs was monitored. Second rosette leaves from 21- or 23-day-old plants were used. The images when the vacuolar membrane engulfs an RCB (25.4–30.5 s in A and 52.6–57.4 s in B) and when an RCB is incorporated into the vacuolar lumen (47.0–52.0 s in A and 65.3–72.9 s in B) are shown. The images in (A) and (B) are still frames from Videos 10 and 11, respectively. Time scales above the images indicate the elapsed time from the start of the respective videos. Open arrowheads indicate an RCB engulfed by the vacuolar membrane. Closed arrowheads indicate the open site of the vacuolar membrane for the release of an RCB into vacuolar lumen. V indicates the region of the vacuolar lumen. Green, VHP1-mGFP; magenta, RBCS-mRFP. Scale bars, 5 µm.

Figure 4—figure supplement 1. Additional observations of the incorporation of Rubisco-containing bodies (RCBs) into the vacuolar lumen.

Additional time-lapse data of the experiments described in Figure 4. The second rosette leaves from 21- (A) or 24-day-old (B, C) plants were used. The images in (A), (B) and (C) are still frames from Videos 12–14, respectively. Time scales above the images indicate the elapsed time from the start of the respective videos. Open arrowheads indicate an RCB engulfed by the vacuolar membrane. Closed arrowheads indicate the vacuolar membrane structure that engulfs an RCB. V indicates the region of the vacuolar lumen. Green, VHP1-mGFP; magenta, RBCS-mRFP. Scale bars, 5 µm.