Chapter title |
Isolation of Autolysosomes from Tobacco BY-2 Cells.
|
---|---|
Chapter number | 12 |
Book title |
Isolation of Plant Organelles and Structures
|
Published in |
Methods in molecular biology, January 2017
|
DOI | 10.1007/978-1-4939-6533-5_12 |
Pubmed ID | |
Book ISBNs |
978-1-4939-6531-1, 978-1-4939-6533-5
|
Authors |
Chihiro Takatsuka, Yuko Inoue-Aono, Yuji Moriyasu |
Editors |
Nicolas L. Taylor, A. Harvey Millar |
Abstract |
Autolysosomes are organelles that sequester and degrade a portion of the cytoplasm during autophagy. Although autophagosomes are short lived compared to other organelles such as mitochondria, plastids, and peroxisomes, many autolysosomes accumulate in tobacco BY-2 cells cultured under sucrose starvation conditions in the presence of a cysteine protease inhibitor. We here describe our methodology for isolating autolysosomes from BY-2 cells by conventional cell fractionation using a Percoll gradient. The autolysosome fraction separates clearly from fractions containing mitochondria and peroxisomes. It contains acid phosphatase, vacuolar H(+)-ATPase, and protease activity. Electron micrographs show that the fraction contains partially degraded cytoplasm seen in autolysosomes before isolation although an autolysosome structure is only partially preserved. |
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Demographic breakdown
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Other | 2 | 4% |
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Unknown | 14 | 30% |
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