Anthocyanin Vacuolar Inclusions Form by a Microautophagy Mechanism.
Anthocyanins are flavonoid pigments synthesized in the cytoplasm and stored inside vacuoles. Many plant species accumulate densely packed, 3- to 10-μm diameter anthocyanin deposits called anthocyanin vacuolar inclusions (AVIs). Despite their conspicuousness and importance in organ coloration, the origin and nature of AVIs have remained controversial for decades. We analyzed AVI formation in cotyledons of different Arabidopsis thaliana genotypes grown under anthocyanin inductive conditions and in purple petals of lisianthus (Eustoma grandiorum). We found that cytoplasmic anthocyanin aggregates in close contact with the vacuolar surface are directly engulfed by the vacuolar membrane in a process reminiscent of microautophagy. The engulfed anthocyanin aggregates are surrounded by a single membrane derived from the tonoplast and eventually become free in the vacuolar lumen like an autophagic body. Neither endosomal/prevacuolar trafficking nor the autophagy ATG5 protein is involved in the formation of AVIs. In Arabidopsis, formation of AVIs is promoted by both an increase in cyanidin 3-O-glucoside derivatives and by depletion of the glutathione S-transferase TT19. We hypothesize that this novel microautophagy mechanism also mediates the transport of other flavonoid aggregates into the vacuole.
Department of Botany, University of Wisconsin-Madison, Madison, Wisconsin 53706 Laboratory of Molecular and Cellular Biology, University of Wisconsin-Madison, Madison, Wisconsin 53706.
Alexandra
A
Chanoca
Center for Applied Plant Sciences, Department of Molecular Genetics and Department of Horticulture and Crop Science, The Ohio State University, Columbus, Ohio 43210.
Nik
http://orcid.org/0000-0002-9396-6626
N
Kovinich
Laboratory for Optical and Computational Instrumentation, University of Wisconsin-Madison, Madison, Wisconsin 53706.
Brian
http://orcid.org/0000-0002-4170-3808
B
Burkel
Department of Botany, University of Wisconsin-Madison, Madison, Wisconsin 53706 Laboratory of Molecular and Cellular Biology, University of Wisconsin-Madison, Madison, Wisconsin 53706.
Samantha
http://orcid.org/0000-0002-6704-5647
S
Stecha
Center for Applied Plant Sciences, Department of Molecular Genetics and Department of Horticulture and Crop Science, The Ohio State University, Columbus, Ohio 43210.
Andres
http://orcid.org/0000-0001-8961-9173
A
Bohorquez-Restrepo
Department of Biological Sciences, University of Tokyo, Hongo, Bunkyo-ku Tokyo 113-0033, Japan.
Takashi
T
Ueda
Laboratory for Optical and Computational Instrumentation, University of Wisconsin-Madison, Madison, Wisconsin 53706.
Kevin W
http://orcid.org/0000-0001-8678-670X
KW
Eliceiri
Center for Applied Plant Sciences, Department of Molecular Genetics and Department of Horticulture and Crop Science, The Ohio State University, Columbus, Ohio 43210.
Erich
E
Grotewold
Department of Botany, University of Wisconsin-Madison, Madison, Wisconsin 53706 Laboratory of Molecular and Cellular Biology, University of Wisconsin-Madison, Madison, Wisconsin 53706 Department of Genetics, University of Wisconsin-Madison, Madison, Wisconsin 53706 otegui@wisc.edu.
Marisa S
http://orcid.org/0000-0003-4699-6950
MS
Otegui
D000872:Anthocyanins / Q000378:metabolism*; D017360:Arabidopsis / Q000166:cytology* / Q000235:genetics / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism; D001343:Autophagy / Q000502:physiology*; D001692:Biological Transport; D003593:Cytoplasm / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005453:Fluorescence; D029965:Gentianaceae / Q000166:cytology* / Q000378:metabolism; D005960:Glucosides / Q000378:metabolism; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D007425:Intracellular Membranes / Q000378:metabolism; D009154:Mutation; D059828:Plant Cells / Q000378:metabolism; D030821:Plants, Genetically Modified; D036226:Seedlings / Q000166:cytology / Q000235:genetics; D014617:Vacuoles / Q000378:metabolism*
D000872:Anthocyanins; D029681:Arabidopsis Proteins; D005960:Glucosides; C462279:cyanidin-3-O-beta-glucopyranoside; D005982:Glutathione Transferase; C484247:TRANSPARENT TESTA 19 protein, Arabidopsis