Abstract
When a fusion protein of cytochrome b5 (Cyt b5) and the red fluorescent protein (RFP) are expressed in tobacco BY-2 cells, the expressed protein forms intracellular aggregates that emit red fluorescence. When such cells are grown to the stationary phase or incubated in nutrient limited medium, RFP fluorescence can be detected in the vacuolar lumen. We investigated this transport mechanism using a limited-nitrogen model. E-64 and 3-methyladenine, which inhibit autophagic processes, blocked the transport of the RFP signal to the vacuole. We next traced the autophagic process in tobacco cells using YFP fused with the tobacco Atg8 homologue (YFP-NtAtg8) and analyzed the contribution of autophagy to the vacuolar transport of the aggregates. Under limited-nitrogen conditions, the aggregates were degraded in preference to other organelles, and the autophagosomes colocalized with the aggregates at a higher frequency than with mitochondria. This is the first demonstration that selective macroautophagic degradation can occur in plant cells.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adenine / analogs & derivatives
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Adenine / pharmacology
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Autophagy / drug effects
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Autophagy / physiology*
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Autophagy-Related Protein 8 Family
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Cell Line
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Cytochromes b5 / genetics
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Cytochromes b5 / metabolism*
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Leucine / analogs & derivatives
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Leucine / pharmacology
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Luminescent Proteins / genetics
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Luminescent Proteins / metabolism
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Microscopy, Immunoelectron
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Microtubule-Associated Proteins / genetics
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Mitochondria / physiology
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Nicotiana / physiology*
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Nitrogen / metabolism*
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Plant Proteins / genetics
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Plant Proteins / metabolism
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Protein Transport / drug effects
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Saccharomyces cerevisiae Proteins / genetics
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Starvation*
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Vacuoles / physiology*
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Vacuoles / ultrastructure
Substances
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ATG8 protein, S cerevisiae
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Autophagy-Related Protein 8 Family
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Luminescent Proteins
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Microtubule-Associated Proteins
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Plant Proteins
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Recombinant Fusion Proteins
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Saccharomyces cerevisiae Proteins
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3-methyladenine
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Cytochromes b5
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Leucine
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Adenine
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Nitrogen
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E 64