Fig. 9: Autophagosome degradation at the petal base during petal abscission is delayed in JA mutants.

Spatiotemporal observation of the autophagosome marker GFP–ATG8a in petal bases of the WT (a) and coi1 (b) by confocal microscopy. Scale bars = 25 µm. c, d Higher magnification images of 35S_pro:GFP–ATG8a in petal bases of the WT (left) and coi1 (right) by confocal microscopy. Images in d are magnified images of the boxed regions in c. The punctate structures labeled by green fluorescence from the cleavage of GFP–ATG8a indicate autophagosome-related structures. Scale bars in c = 10 µm. Scale bars in d = 1 µm. Numbers in the right corners indicate flower position. e GFP–ATG8a fluorescence-intensity profile in WT and coi1 autophagosome-like structures. n = 9. Each gray line indicates an individual trace. The green line shows mean data. f Immunoblot analysis showing the processing of GFP–ATG8a in the WT and the coi1 mutant during petal abscission. Crude protein extracts from petals at the indicated positions were subjected to SDS–PAGE and immunoblot analysis with anti-GFP antibodies. Arrows and arrowheads to the right side of immunoblot image indicate GFP–ATG8a and free GFP, respectively. Histone H3 served as a loading control. g Quantification of the GFP/GFP–ATG8a ratio during petal abscission by densitometric scans of the immunoblots shown in g. Values are means ± SE (n = 3 independent experiments). Asterisks indicate statistically significant differences between the WT and the coi1 mutant based on two-tailed Student’s t test *p = 0.04.